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Laboratory Information and Courses from MediaLab, Inc.

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Laboratories Individuals

Alpha Thalassemia
Silent Carrier

The Silent Carrier form of alpha thalassemia results from one alpha chain loci deletion. Individuals who are silent carriers show no clinical disease and demonstrate normal results during routine laboratory testing. This form of alpha thalassemia is usually discovered upon family studies.

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References

Burtis, CA. & Ashwood, ER. Tietz Textbook of Clinical Chemistry 2nd ed. W. B. Saunders. 1994.Harmening, DM. Clinical Hematology and Fundamentals of Hemostatis 5th ed., F.A. Davis, 2008Lotspeich-Steininger, Stiene-Martin and Koepke, Clinical Hematology Principles, Procedures, Correlations, Lippincott 1992McKenzie, SB., Textbook of Hematology 2nd ed., Williams and Wilkins 1996.Miale, JB, Laboratory Medicine Hematology 6th ed., Mosby 1982.Nouwens, J and Spahn, M. Hemoglobin H Disease: A self-instructional unit 3rd ed., Educational Materials for Health Professionals, Inc. 1991.Doig, K. Rodak's Diagnostic Hematology 3rd ed. W.B.Sunders Co., 2007.

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What laboratory tests should be performed to aid in the diagnosis of this anemia?View Page

Antibody Detection and Identification
References

Alba MA. Clinical Immunohematology Laboratory Manual. Albuquerque, NM: UNM Health Sciences Center; 2008.Brecher MF, Leger RM, Linden JV, Roseff SD, eds. Technical Manual 15th ed. Bethesda, Md. AABB; 2005.Harmening DM. Modern Blood Banking and Transfusion Practices. 5th ed. Philadelphia, Pa: F.A. Davis Company; 2005.

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Examples of Antibodies to Low-Incidence Antigens

Antibodies to low-incidence antigens will be difficult to test for since most screen and panel cells do not have these antigens on the testing cells. Further testing may be needed at a reference laboratory where a larger selection of antibody panels are available to locate cells positive for these antigens.Suspect an antibody to a low-incidence antigen if: AHG crossmatch is incompatible and Other causes have been ruled out (positive donor DAT, ABO incompatibility) Examples of antibodies to low-incidence antigens are: anti-V, anti-Cw, anti-Kpa, anti-Jsa, and anti-Lua.

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Beta Thalassemia
References

Burtis CA, Ashwood ER. Tietz Textbook of Clinical Chemistry 2nd ed. WB Saunders; 1994.Doig, K. Rodak's Diagnostic Hematology. 3rd ed. WB Saunders Co; 2007.Harmening DM. Clinical Hematology and Fundamentals of Hemostatis. 5th ed. FA Davis; 2008Hoffman R, Benz EJ Jr., Shattil SJ, Furie B, Cohen HJ, Silberstein LE. Hematology Basic Principles and Practice, 2nd ed. Churchill Livingstone; 1995.McKenzie SB. Textbook of Hematology, 2nd ed. Williams and Wilkins; 1996.Miale JB, Laboratory Medicine Hematology, 6th ed. Mosby; 1982.Stiene-Martin EA, Lotspeich-Steininger CA, Koepke JA, Clinical Hematology Principles, Procedures, Correlations, 2nd ed. Lippincott; 1998.

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Laboratory Test Results

Test Patient Result Reference Intervals (Adult female) White blood cell (WBC) count 3.7 x 109/L 4.4 - 11.3 x 109/L Red blood cell (RBC) count 5.6 x 1012/L 4.1 - 5.1 x 1012/L Hemoglobin (Hb) 10.5 g/dL 12.3 - 15.3 g/dL Hematocrit (HCT) 36.6% 35.9 - 44.6% MCV 65.8 fL 80.0 - 96.0 fL MCH 19.9 pg 27.5 - 33.2 pg MCHC 26.7% 33.4 - 35.5% RDW 14.0 <14.5 Platelets 249.0 x 109/L 100.0 - 450.0 x 109/L Total serum iron 165 µg/dL 60 - 150 µg/dL Iron-binding capacity 230 µg/dL 250 - 400 µg/dL The RBC count is increased for the amount of hemoglobin present. The concentration of hemoglobin in the RBCs is slightly decreased (hypochromic) and the cells are small (microcytic). The variation in RBC size (RDW) is within normal limits.

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Case History Summary

The laboratory findings in this case represent classic findings seen in beta thalassemia minor including: erythrocytosis, decreased hemoglobin, normal hematocrit, normal RDW, and the presence of codocytes (target cells). This patient does have a mild anemia, but some patients with beta thalassemia minor have no anemia. Hemoglobin electrophoresis confirms this diagnosis, showing an increased Hb A2 level and decreased Hb A.In addition, the slightly increased iron and slightly decreased TIBC contradict a suspicion of iron deficiency. These chemistry results are typical for beta thalassemia, even though the red blood cells are microcytic and hypochromic.

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Cerebrospinal Fluid
Specimen Labeling and Transport

The cerebrospinal fluid sample should be properly labeled with the tube number, patient's name and hospital number. The samples should be transported to the laboratory immediately.

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Safety Precautions

Important safety precautions must be observed when handling cerebrospinal fluid. The following guidelines apply:Semi-automatic micropipettes and disposable plastic chambers are the safest option for CSF testing. Many laboratories still use the hemacytometer with disposable pipets.If disposable materials are not used, soak contaminated reusable pipets, hemacytometer and coverslip in 70% alcohol or Wexide.All disposable items should be placed in a biohazard container for appropriate disposal.Wash hands thoroughly when the examination is completed.Spinal fluids which are to be discarded must be placed in biohazard containers for appropriate disposal.Careful attention to specimen processing and handling will help ensure that accurate results are obtained.

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Normal Characteristics

Normal cerebrospinal fluid has the following characteristics:colorlessclearno clot presentspecific gravity of 1.006 - 1.008pH 7.3When the specimen is received in the laboratory, the macroscopic examination is performed immediately. The specific gravity examination may be optional in some laboratories.

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WBC Correction for Traumatic Tap

A calculation is used to correct CSF WBC counts which are falsely increased due to a traumatic tap: WBCs added = WBC(blood) x RBC(CSF) / RBC(blood)The blood WBC count is multiplied by the ratio of the cerebrospinal fluid RBC count to blood RBC count.The result is the number of artificially introduced WBCs. The true CSF white cell count is then calculated by subtracting the artificially introduced WBCs from the actual CSF WBC count. If the patient's peripheral WBC and RBC counts are within normal limits, some laboratories use the following formula: Subtract one white cell from the CSF WBC count for each 750 RBC counted in the spinal fluid.

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Cytospin Technique

In the cytospin procedure, use a high speed centrifuge to concentrate the cells on a slide in a uniform monolayer 6 mm in diameter. The monolayer distribution enhances the morphological appearance of the cells present.Allow the slides to dry in air for several minutes and then stain them with Wright-Giemsa stain. Cytospin slides may be placed in an automatic stainer, such as Hema-Tek, or stained manually.Perform a 100 or 200 cell differential and record the number of neutrophils, eosinophils, basophils, lymphocytes, monocytes, macrophages, and blasts cells.Pathologists must review any slide which has tumor cells, unidentified cells, or immature stages of cells, such as blasts.Since criteria for review may vary from one laboratory to another, be sure to check the requirements in your laboratory before reporting the differential.

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Chemical Screening of Urine by Reagent Strip
A voided urine specimen is delivered from the women's clinic to the laboratory six hours after collection. The following results are reported: Color: yellowProtein: negativeBilirubin: negative Clarity: cloudyGlucose: negativeUrobilinogen: 0.2 mg/dL Sp. Gravity: 1.020Ketone: negativeNitrite: positive pH: 9.0Blood: negativeLeukocyte esterase: negativeWhat might these results indicate?View Page
Precautions

The reagent strips must be handled and stored properly in order to ensure that results are accurate. The following precautions should be observed: Store strips according to the manufacturer's recommendation. DO NOT expose strips to moisture, direct sunlight or volatile fumes. Remove only enough strips for immediate use and immediately recap the bottle. Avoid contamination of test strips. Do not touch the test areas with fingers or do not lay the test strips directly on the workbench. DO NOT use discolored strips. Compare the color of the unused strip to the negative area on the color chart provided by the company. The color should be similar. Check the expiration date. Re-label the container with a revised expiration date if the manufacturer states a shortened usage period once the container has been opened. Reagent strips must be tested periodically (frequency defined by the laboratory) for clinical reactivity with normal and abnormal urine controls. Urine controls are available commercially or may be prepared and preserved in-house.

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Procedure Caution

Although the procedure is simple to perform, accurate results depend on careful adherence to manufacturer’s directions and adequate quality control. Normal and abnormal controls should be tested whenever a new lot of strips is opened, and at the frequency defined by the laboratory's procedure. If quality control results do not correspond to the published control values, the problem must be resolved before patient samples are tested. High levels of ascorbic acid (Vitamin C) in the urine may inhibit some reagent strip reactions, such as glucose, blood, bilirubin, nitrate and leukocyte esterase. The urine dipstick's package insert will provide information about potential interfering substances, including ascorbic acid. Intensely colored urine may make it difficult to correctly interpret color reactions on the dipstick. The affected tests should not be reported from the dipstick. It would be necessary to use an alternative method of testing if available.

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Quality control procedures should be performed with each new lot of reagent strips, and as often as required by the laboratory's procedure.View Page
A urine specimen was collected at 6:00 A.M. and remained at room temperature until it was received in the laboratory at 3:30 P.M. How may the pH of the specimen be affected by the extended time at room temperature if bacteria are present in the specimen?View Page
Confirmatory Testing for Protein

Semiquantitative tests are used in some laboratories to confirm the presence of protein in the specimen when the result is positive on the urine dipstick. Tests that are used for confirmation include: sulfosalicylic acid (SSA); heat and acetic acid; nitric acid ring test; and Roberts' Ring Test. Any one of these procedures may be used for confirmation of the presence of protein. A protein dipstick result that is greater than a trace may be an indication of proteinuria.

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False Negative Results

False negative results occur when elements present in the urine interfere with either the enzymatic reaction or prevent the oxidation of potassium iodide. Examples of such substances include: large quantities of ketones aspirin ascorbic acid > 50 mg/dL with some reagent strips levadopa 5-hydroxyindoleacetic acid homogentisic acid sodium fluoride ( a preservative)A specific gravity higher than 1.020 may lower glucose reagent sensitivity, especially in the presence of a high urine pH. Exposing reagent strips to excess humidity may also reduce glucose reagent reactivity.Check the package insert of the reagent strips used in your laboratory for interfering substances that may affect glucose results.

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Test for Reducing Substances Other than Glucose

Urine specimens from certain pediatric patients should be tested for other reducing substances, such as galactose, when the results for glucose are negative using the routine dipstick method. The laboratory's procedure should define when additional testing is needed.

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Semi-Automated Instruments

Several manufacturers offer semi-automated instruments (dipstick readers) for reading reagent strips. Use of an instrument removes the subjectivity of visually interpreting color changes on reagent strips, and assures that tests will be read at the correct time. Transcription errors will also be avoided if the instrument is interfaced with the laboratory information system. The technology employed is based on the principle of reflectance, with the amount of light reflected being inversely related to the concentration of substances present. An example of reflectance is the light which is scattered after light strikes an unpolished surface. Since each component on the dipstick produces a different color reaction, the light source for each test must be at the appropriate wavelength. This is accomplished either by using filters or monochromatic light sources. The percent reflectance is determined by dividing the test reflectance by the calibration reflectance and multiplying by 100. Algorithms are used to change the results obtained into a linear relationship with concentration of analyte.

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CLIA General Laboratory Review
Which one of the following does not directly regulate clinical laboratories:View Page
Which of the following infectious agents represent the greatest risk to the laboratory worker:View Page
A laboratory fire that is the direct result of the electrical malfunction of a laboratory instrument or piece of equipment would be classified as:View Page
If a laboratory 's control range (using a 99.7 confidence interval) for a given assay is 20.0 to 50.0, what would its means and one standard deviation be:View Page
Many laboratory procedures are conducted at 37o C. This corresponds to what temperature on the Fahrenheit scale:View Page
What minimum level of specific resistance (megohms@25o C) is required for a Type I water system:View Page
What percentage solution of sodium hypochlorite (bleach) is recommended as a routine laboratory disinfectant:View Page
Which of the following microscopic techniques is capable of producing a 3-dimensional image :View Page
CPT 4 codes:View Page
Which of the following is not directly responsible for setting and monitoring competency requirements for laboratory personnel?View Page
Which of the following is the main function of the ASCP Board of Registry:View Page

CLIA Hematology / Hemostasis Review
When three tubes of cerebrospinal fluid are received in the laboratory they should be distributed to the various laboratory sections as follows:View Page

CLIA Microbiology / Serology Review
Which of the following statements about Rickettsia is false:View Page

Confirmatory and Secondary Urinalysis Screening Tests
Screening and Secondary Tests for Protein

A routine reagent strip protein method, based on the principle of "protein error of indicators," produces a visible colorimetric reaction that is capable of detecting most instances of proteinuria.Traditionally, laboratories have used sulfosalicylic acid (SSA) to confirm all positive protein reagent strip results, but this practice is not as common in today's laboratories. SSA is a precipitation method that reacts with all forms of protein. However, any substance that is precipitated by acid will produce false-positive SSA results. This includes radiographic dyes, cephalosporins, penicillins, and sulfonamides. SSA may be used as a secondary protein detection method if the urine is highly alkaline (pH of 9.0 or greater) which would overwhelm the buffering capacity of the reagent on the protein reagent stick. SSA may also be used as an alternative protein detection method if the urine is highly colored so that the colorimetric reaction is masked on the reagent strip.

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A urine specimen to be tested for bilirubin arrives in the laboratory after sitting on the counter at the nurses station for 2 hours. Which of the following statements describes the Ictotest® reaction that could potentially occur in this situation?View Page
Limitations of the Procedure

The product profile for Ictotest® points out that bilirubin is very light sensitive, so urine specimens should be protected from excessive light exposure and examined as quickly as possible when received in the laboratory. On standing, bilirubin, which has a goldish color, is oxidized to biliverdin, which is a green color. Many of the procedures used to detect bilirubin will not react with biliverdin, so false-negative results may occur if urine is not fresh when tested.

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Testing for Reducing Substances Other Than Glucose

Testing pediatric urine specimens for reducing substances other than glucose is a policy that should be implemented in the urinalysis laboratory. The maximum age for this testing is defined by each laboratory and is usually based on consultation with the pediatric clinical staff. The policy that is implemented in most laboratories is to test urine specimens for other reducing substances if the glucose test on the reagent strip is negative and the urine specimen is from a child below the age of one. Verify the policy for your own laboratory because the cutoff age for testing may be different.

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The 2-Drop Clinitest® Procedure

Record the test result that is associated with the color block that most closely matches the color of the test in the tube. Remember that the final color may not be the test result if the "pass through" phenomenon occurred. Test results should be recorded according to your laboratory's procedure. Laboratories may choose to record results as 1%, 2%, 3%, etc; 1+, 2+, 3+, etc; or 100mg/dL, 200mg/dL, 300mg/dL, etc. The result of the test in the top image is negative, and the result of the test in the bottom image, reported as a percentage, is 2%. (Note: Colors in the photograph may vary slightly from actual test colors.)

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Current Topics in Clinical Microbiology
Review 2

Smith KR, Fisher HC III, Hook, EW III: Prevalence of fluorescent monoclonal antibody-nonreactive Neisseria gonorrhoeae in five North American sexually transmitted disease clinics.J Clin Microbiol 34:1551-1552, 1996We compared a direct fluorescent monoclonal antibody (DFA) test with alternative enzymatic and fermention tests for identifying presumptive gonococcal isolates in a systematic sample from patients attending five sexually transmitted disease clinics in five cities.Fourteen (2.5%) of 556 isolates from three clinics were nonreactive with the DFA confirmatory reagent and reactive by both the Quad-Ferm and Rapid NH tests. The prevalence of DFA-nonreactive Neisseria gonorrhoeae isolates varies geographically and is independent of local methods for the identification of possible gonococci.On the basis of our findings, we recommend that for use in medicolegal and other instances in which a diagnosis of gonorrhea has the potential to have far-reaching effects, it is appropriate to test DFA reagent-nonreactive, oxidase-positive, gram-negative diplococci by alternative methods of gonococcal confirmation.Although the prevalence of such isolates could change, the fluorescent monoclonal antibody confirmation reagents remain useful for many clinical situations. Their ease of use and ready applicability for screening large numbers of isolates make them useful for many laboratories.

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Acute Onset Pneumonia

A 70-year-old transient with productive cough, pleuritic chest pain radiating to the mid back, fever, and chills was seen in the emergency room. Expectorated sputum was sent to the laboratory for gram stain and culture. (Continue on next page)

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The oxacillin screen test alone is not sufficient for determining the susceptibility to penicillin for S. pneumoniae isolates recovered from cerebrospinal fluid (CSF).View Page
Clinical isolates of Escherichia coli and Klebsiella pneumoniae may possess ESBL activity. Therefore, clinical laboratories should be screening all clinically significant isolates of these two species.View Page
Clinical History

A 72- year old woman had a history of recurrent urinary tract infections over the past several months, for which she had received different regimens of antibiotics including ampicillin, trimethoprim-sulfasoxazole, and ciprofloxacin.Relapses often occurred 10 days to two weeks after cessation of therapy.The current flare up, manifest by dysuria, lower abdominal pain and cloudy urine was accompanied by shaking chills and spiking fever.A sterile mid-stream urine specimen was sent to the laboratory for culture.

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Enterococcus faecium ID

As a high percentage of Enterococcus faecium strains carry the Van A gene and are highly resistant to vancomycin. Species identifications are performed in some laboratories where MIC susceptibility testing may not be available.Methods for the phenotypic separation of E. faecium from E. faecalis are limited.Illustrated in this photograph are positive reactions for acid production from arabinose and melibiose (yellow color), characteristic of E. faecium. E. faecalis are negative for these reactions.A few preformed substrates such as beta galactosidase (E. faecium positive, E. faecalis negative) also serve to separate these two species, accomplished by certain commercial systems that include these substrates.E. faecium is not motile, an additional characteristic helpful to separate vancomycin-resistant Enterococcus species from E. cassiloflavus and E. gallinarum, both of which are motile, and carry the low level resistant gene VAN-c.

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Vancomycin Resistance

Vancomycin and ampicillin resistance among Enterococcus species, particularly E. faecium have been on a steady increase.The disk diffusion screening test is used in many laboratories to detect vancomycin resistant strains. Note in the upper photograph that no zone of inhibition is seen around either the vancomycin or the ampicillin disk, indicating resistance to both drugs.Vancomycin-resistant Enterococci (VRE) have been divided into three phenotypes--Van A, Van B, and Van C.Vancomycin-resistant strains of E. faecalis and E. faecium are commonly of the Van A phenotype, demonstrating high level resistance (MIC's higher than 64 ug/mL), as illustrated by total resistance of the test strain in the E test and the VA disk, as illustrated in the lower photograph.The strain shown in the lower photograph, however, is ampicillin susceptible at the level of 1 ug/ml (see lower set of yellow arrows), indicating that this drug may be effective in treating the urinary tract infection.

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Review 2

Citron DM. Appelbaum PC.: How far should a clinical laboratory go in identifying anaerobic isolates, and who should pay? Clinical Infectious Diseases. 16 Suppl 4:S435-8, 1993Identification of anaerobic bacteria in specimens from sites of infection due to mixed organisms can be time-consuming and expensive. Laboratories should limit anaerobic workups by testing only those specimens that have been properly collected and transported to the laboratory.Use of selective and differential media for initial processing can provide rapid and relevant information to the clinician. Anaerobes isolated from normally sterile sites and sites of serious infection should always be completely identified. Group-or genus-level identifications may suffice in other instances.The Bacteroides fragilis group of organisms should always be identified because of their virulence and resistance to many antimicrobial agents.Some of the other organisms that warrant identification include Clostridium septicum (associated with gastrointestinal malignancy); Clostridium ramosum, Clostridium innocuum, and Clostridium clostridioforme (which are resistant to antibiotics); Clostridium perfringens (a cause of myonecrosis and gas gangrene,potentially serious infection); anaerobic cocci (which may be resistant to metronidazole and clindamycin); and fusobacteria (which may be virulent and resistant to clindamycin and penicillin).

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MRSA Screen

Perhaps the most efficient means for detecting methicillin-resistant staphylococci in clinical laboratories is the use of the agar dilution screening test.Illustrated in the photograph is a Mueller-Hinton agar plate containing 6ug/ml of oxicillin, previously inoculated with a strain of Staphylococcus aureus. Oxacillin is used as a marker for methicillin resistance because it is more stable in the agar medium. Growth on this screening medium is presumptive for methicillin resistance.Thus, in the presence of growth, as shown here, a follow-up MIC test must be performed to determine the exact level of resistance.

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Most strains of S. milleri (anginosus) carry the F antigen (see photograph). Rare strains that carry the group A antigen can be differentiated from S. pyogenes by which of the following laboratory tests:View Page
Clinical History

The prototype history for this organism is either a still birth or a neonate with death ensuing within 2 or 3 days post-partem due to high fever, sepsis, and respiratory distress. The mother usually experienced a flu-like illness late in the third trimester of pregnancy, characterized by low-grade fever, myalgias, malaise and backache. In this case, biopsy material of brain tissue obtained at autopsy was submitted to the pathology laboratory for tissue diagnosis and fluid from the pia-arachnoid was sent to the microbiology laboratory for culture.

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Cellulitis Lesion

A 35 year old man presented in the emergency room with an erythematous, vesiculo-pustular lesion of the arm near the elbow (see photograph). One week previously he had scratched his arm on the aerial of his car while washing the windshield. He noticed a red area about 3 days after the incident, which then spread to involve the adjacent tissue. The central pustule developed on the day he was seen. Material from the center of the pustule was sent to the microbiology laboratory for culture.

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Department of Transportation (DOT) & Federally Regulated Urine Specimen Collection Training
Custody and Control Form

While the Federally Regulated CCF must be used exclusively for DOT drug screen collections, there are extenuating circumstances when another form of CCF may be used. For example, where a post-accident collection must be made and the collector does not have time to obtain a Federally Regulated CCF. In this situation, a Non-Federally Regulated CCF may be used. The collector should note in the "Remarks" section why the Non-Regulated CCF was used.The use of a Non-Regulated CCF for a Federally Regulated collection is not a reason for the laboratory to refuse to test the specimens nor the Medical Review Officer (MRO) to release the results. The use of a Non-Regulated CCF for a Federally Regulated drug screen is a "correctable flaw," which may include the collector detailing the reason for using a Non-Federally Regulated CCF in a "Memorandum for Record."

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Donor completes certification statement

The collector now directs the donor to read, sign, and date the certification statement located on the "pink" page of the control form. The donor must provide date of birth, printed name, and day and evening contact telephone numbers.The collector completes the collector's portion of the chain of custody on the control form by printing his or her name, signing where indicated, and recording the date and time of the collection. The collector must also enter the specific name of the delivery or courier service transferring the specimens to the laboratory if applicable.

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Observed collection scenarios

Scenario 4: The donor returns from the restroom with a sufficient specimen. It is very warm to the touch. The collector is unable to obtain a reading from the temperature strip. Collector’s response: The collector completes the collection and prepares the specimen for shipment. The collector explains the situation with a supervisor. If the supervisor concurs that an observed collection is in order, the collector next tells the donor that a new collection will be conducted under direct observation. The collector explains that because the temperature of the specimen was not within the acceptable range (90-100º F/32-38º C) there is suspicion of substitution or adulteration. A new CCF is initiated. The collector marks on the CCF that the collection is observed and notes under Remarks why it is observed. The collector also notes the control number of the suspect collection. The observed specimen along with the suspect specimen are both shipped to the laboratory in separate plastic tamper-resistant bags.

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Broken Security Seal

If a security seal is broken while being removed from the CCF or during the application of the first seal on the primary specimen vial, the collector should transfer the information to a new CCF and use the seals from the second CCF.If one seal is already in place on a specimen vial and second seal is broken while being removed from the CCF or is broken during application on the second specimen vial or while the employee is initialing either seal, the collector should initiate a new CCF and note in the "Remarks" section how the seals were broken. The seals from the second CCF should be placed perpendicular to the original seals to avoid obscuring information on the original seals. The donor must initial the second set of seals also. The initials on all the seals must match. The collector should then draw a line through the specimen ID number (and bar code if present) on the original seals to ensure that the laboratory does not use that number for reporting the results. The collector must not pour the specimen into new vials.

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Observed collection scenarios

Scenario 5: The collector notices that the urine the donor just handed to her has a very strong smell like that of a cleaning product such as bleach. Collector’s response: The collector completes the collection in the usual manner and prepares the specimen for shipment. The collector explains the situation to a supervisor. If the supervisor concurs that an observed specimen should be collected, the collector explains to the donor that because of the strong, unusual smell, the first specimen is suspect for adulteration and that a directly observed collection will be done. A new CCF is initiated. The collector marks on the CCF that the collection is observed and notes under Remarks why it is observed. The collector also notes the control number of the suspect collection. The observed specimen along with the suspect specimen are both shipped to the laboratory in separate plastic tamper-resistant bags. In addition to an unusual smell, other indications of adulteration might be an unusual color that cannot be explained by medication, particles or debris in the urine, and a heavy or thick foam that is inconsistent with urine.

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Fatal Flaws and Correctable Flaws

Fatal FlawsIt is important to remember that the following are fatal flaws and can cause the specimen not to be tested: Number on Custody and Control Form and security strips do not match. Security strip on the specimen vial is broken or shows evidence of tampering. Quantity of urine needed is not sufficient. There is no printed collector's name or signature.Correctable FlawsThe following are flaws that may be corrected by either sending a signed statement or a Memorandum for Record to the laboratory: The collector printed his or her name, but forgot to sign the CCF. The collector checked the temperature of the specimen, but forgot to note this fact on the CCF.

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Descriptive Statistics
Why Statistics?

Many people involved in the clinical laboratory sciences need to be familiar with basic statistics for a variety of reasons.  These reasons include: performing quality control, and interpreting of results of instrument testing determining suitability of different methods or instruments for the same task understanding how acceptable laboratory procedures and methods are established determining ranges for clinical tests of normal, healthy individuals understanding clinical trials and new methods presented in journals and articles performing those trials and research projects yourself

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Electrophoresis
Introduction

Electrophoresis is the migration or separation of charged particles or solutes of a liquid solution in an electrical field. Conventional electrophoresis is tedious and time consuming. Electrophoresis automation and newer electrophoresis techniques have revitalized the utilization of electrophoresis in today's clinical laboratories. Molecular diagnostic analysis using electrophoresis and research in proteomics have also contributed to this revitalization.

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Types of Electrophoresis

There are numerous applications of electrophoresis. Routine protein electrophoresis performed in clinical laboratories is the oldest method and therefore the most frequently used method. With the advent of molecular diagnostics, several other electrophoresis methods have become very important, highly automated, and have several important applications.Types of electrophoresis that will be discussed are Routine electrophoresis High resolution electrophoresis Polyacrylamide gel electrophoresis Capillary electrophoresis Isoelectric focusing Immunochemical electrophoresis Two-dimensional electrophoresis Pulsed field electrophoresis

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References

Clinical Chemistry Concepts and Applications. Shauna C. Anderson and Susan Cockayne. Long Grove, Illinois: Waveland Press, Inc, 2003.Clinical Laboratory Instrumentation and Automation Principles, Applications, and Selection. Kory M. Ward, Craig A. Lehmann, Alan M. Leiken. Philadelphia: WB Saunders Company, 1994.Laboratory Instrumentation, 4th Edition. Mary C. Haven, Gregory A. Tetrault, Jerald R. Schenken, eds. New York: Van Nostrand Reinhold, 1995.Molecular Diagnostics Fundamentals, Methods, and Clinical Applications. Lela Buckingham and Maribeth L. Flaws. Philadelphia: FA Davis Company, 2007.Principles of Gel Electrophoresis. Available at http://www.vivo.colostate.edu/hbooks/genetics/biotech/gels/principles.html accessed 9/29/08.Tietz Textbook of Clinical Chemistry and Molecular Diagnostics, 4th Edition. Carl A. Burtis, Edward R. Ashwood, David E. Burns, eds. Philadelphia: Elsevier Saunders, 2005.

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Automation

Automated systems for protein electrophoresis are available for large volumes of samples for electrophoresis. An automated system is capable of separating 10-100 samples simultaneously. There are several different automated systems and the number of process steps automated varies. Automated steps may include reagent addition, sample application, electrophoresis separation, staining, and detection.Below is a stained electrophoresis land for one sample analyzed on the automated instrument, SPIFE 3000, Helena Laboratories. The separated proteins are stained with Acid Blue, a Helena Laboratories' stain developed from Coomassie dye.

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Immunofixation Electrophoresis

An agarose gel electrophoresis first separates the proteins in a serum sample. Antiserum against the protein of interest is spread directly on the gel. The protein of interest precipitates in the gel matrix. After a wash step to remove other proteins, the precipitated protein is stained. This method is qualitative and is used to identify proteins found in multiple myeloma.Below is the immunofixation electrophoresis gel from a serum sample analyzed on SPIFE 3000, Helena Laboratories. After electrophoresis, the precipitated proteins are stained with Acid Violet, a stain developed and used by Helena Laboratories. The SP lane represents a routine serum protein electrophoresis of this specimen. On the next three protein separations, antiserum against IgG, IgA, and IgM were applied to the G, A, M lanes respectively. Antiserum to kappa light chain was added to the next protein separation and antiserum to lambda light chain to the last protein separation.

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Densitometry

After electrophoresis, a stained gel is passed through the optical system of a densitometer to create an electrophoregram, a visual diagram or graph of the separated bands. A densitometer is a special spectrophotometer that measures light transmitted through a solid sample such as a cleared or transparent but stained gel. Using the optical density measurements, the densitometer represents the bands as peaks. These peaks compose the graph or electrophoregram and are printed on a recorder chart or computer display. Absorbance and/or fluorescence can be measured with densitometry.An integrator or microprocessor evaluates the area under each peak and reports each as a percent of the total sample. If the electrophoresis is for separation of serum proteins, the concentration of each band is derived from this percent and the total protein concentration. If the electrophoresis is for separation of enzymes, the enzyme activity of each band is derived from this percent and the total enzyme activity. The densitometer scan below depicts the separated bands from a serum sample electrophoresis. The SPIFE 3000, Helena Laboratories, electrophoresis splits the beta zone into two fractions for easier detection of small beta-migrating monoclonal gammopathies. The densitometer scan from this electrophoresis shows five bands with two peaks in the beta band.

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Emerging Cardiovascular Risk Markers
Risk Markers

We have listed the 'classic' cardiovascular risk markers as LDL-C, HDL-C and triglycerides. But there are many more cardiovascular risk markers as well as cardiovascular risk factors. A cardiovascular risk factor is a condition (not a laboratory analyte) that is associated with an increased risk of developing cardiovascular disease. Examples include: Age Gender (males are at increased risk) Heredity Hypertension Cigarette Smoking Obesity Diabetes StressThere are also negative risk factors, factors which decrease a person's risk of cardiovascular disease. Examples include: Optimal HDL-C concentration Exercise Estrogen Moderate alcohol intakeThis course will not focus on cardiovascular risk factors. Instead we will focus on newer, emerging cardiovascular risk markers. There are well over twenty well-studied cardiovascular risk markers; in this course we will focus on some of the more established markers and the ones which are becoming more commonly measured in the clinical laboratory. These include apolipoprotein A1/apolipoprotein B100, Lp(a), oxidized LDL, LpPLA2, hsCRP and lipoprotein particle size and concentration.It is important to remember that the association between a cardiovascular risk marker and actually having or developing cardiovascular disease is a statistical one. The fact that a patient has a particular risk marker which is abnormal simply increases the probability of developing cardiovascular disease, it does not mean that he or she is certain to develop cardiovascular disease. Conversely, if an individual does not have a particular cardiovascular risk marker present it does not guarantee protection against cardiovascular disease. We must always remember that some percentage of individuals who have heart attacks or strokes will not have abnormal risk markers present.

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Lp(a) Testing

One of the problems with Lp(a) measurement is that the Apo(a) protein has a variable mass. It can have a molecular weight ranging from 275,000 to 800,000 daltons. This is due to variable amounts of repeating regions of the protein. Immunoassay antibodies which recognize these regions will thus give more signal for larger Apo(a) molecules compared to smaller Apo(a) molecules. This is not ideal since again, we would prefer to quantify the number of particles and Lp(a) containing large Apo(a) molecules will produce more signal, skewing the count. One assay system that tries to correct for this is the Lp(a) Cholesterol Electrophoresis Assay sold by Helena Laboratories. This assay uses electrophoresis followed by cholesterol staining and densitometry to calculate the concentration of cholesterol in Lp(a). Although this method still does not enumerate particles, it does appear to have less heterogeneity.Lp(a) is an acute phase reactant. This means that Lp(a) levels will rise in the context of general inflammation. Thus, Lp(a) should not be measured when there is extensive inflammation, such as immediately following a cardiovascular event. Concentrations of Lp(a) above 30 mg/dL are associated with increased cardiovascular risk. The risk of having a cardiovascular event increases 2 to 3 fold if Lp(a) cholesterol is > 30 mg/dL. Fifteen to 20% of the Caucasian population have Lp(a) levels >30 mg/dL. Africans, or people of Aftican descent, generally have levels higher than Caucasians and Asians, however, results must be evaluated in conjunction with clinical history.

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Summary

In this course we have described some emerging cardiovascular risk markers. It is important to note that there are many more markers, some of which appear robust and which may have clinical value. Examples of other risk markers that are emerging but were not discussed are listed in the table to the right.An important question that should always be asked is "how many risk markers do we need?" With so many risk markers available, the laboratory needs to research which markers are truly the strongest and most valuable for the patient demographic the facility deals with most and which markers physicians will order and utilize.

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Adult Treatment Panel

How do physicians interpret risk marker results? Assuming the laboratory offers, and physicians order, cardiovascular risk marker tests, how are these results used? The National Cholesterol Education Program periodically assembles scientists and physicians to create lipid treatment guidelines for patients. These panels are referred to as the Adult Treatment Panel (ATP). The third assembly of the ATP did not give specific guidelines regarding risk marker use in patients but they did acknowledge their potential utility. The general consensus is that novel cardiovascular risk markers should be used in selected patients, such as those who already have significant risk factors (hypertension, smoking, obesity, etc.) or in patients who have family histories of cardiovascular disease. The value in using risk markers is that they will not only uncover cardiovascular risk but they can also be used to motivate patients to alter lifestyle and diet. It is expected that as these emerging cardiovascular risk markers continue to be validated in clinical studies, they will become very useful and perhaps even be part of a new standard of care for patients.If risk marker levels can be correlated to treatment strategies, physicians will find them especially useful in tracking patient success.

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References

Atherosclerosis. U.S. Department of Health & Human Services National Institutes of Health. Available at http://www.nhlbi.nih.gov/health/dci/Diseases/Atherosclerosis/Atherosclerosis_WhatIs.htmlAccessed June 23, 2009.Daniels LB, Barrett-Connor E, Sarno M, Laughlin GA,Bettencourt R, Wolfert RL. Lipoprotein-associated phospholipase A2 (Lp-PLA2) independently predicts incident coronary heart disease (CHD) in an apparently healthy older population: The Rancho Bernardo study. J Am Coll Cardiol. 2008;51:913-919.Executive Summary of the third report of the National Cholesterol Education Program (NCEP) Expert Panel on Detection, Evaluation, and Treatment of High Blood Cholesterol in Adults (Adult Treatment Panel III). JAMA. 2001; 285:2486-2497. Frostegard, J, Wu R, Lemne C, Thulin T, Witztum JL and de Faire U. Circulating oxidized low-density lipoprotein is increased in hypertension, Clin Sci 2003; 105, 615.Garza CA, Montoir VM, McConnell JP, et al. Association between lipoprotein-associated phospholipase A2 and cardiovascular disease: a systematic review. Mayo Clin Proc. 2007;82(2):159-165.Interpretive Handbook, (MC0440rev0407) Mayo Clinic, Rochester MN;2007. Maksimowicz-McKinnon K, Bhatt DL, Calabrese LH: Recent advances in vascular inflammation: C-reactive protein and other inflammatory biomarkers. Curr Opin Rheumatol. 2004;16:18-24.Mora S, Szklo M, Otvos JD, et al. LDL particle subclasses, LDL particle size, and carotid atherosclerosis in the multi-ethnic study of atherosclerosis. Atherosclerosis. 2007;192:211-217.NACB Laboratory Medicine Practice Guidelines. Emerging biomarkers of cardiovascular disease and stroke. National Academy of Clinical Biochemistry Laboratory Medicine Practice Guidelines. 2006.PLACtest animation, diaDexus. http://www.plactest.com/laboratorians/action.php Accessed June 23, 2009.Rifai N, Warnick GR. Lipids, lipoproteins, apolipoproteins, and other cardiovascular risk factors. In: Burtis CA, Ashwood ER. Bruns DE. Tietz Textbook of Clinical Chemistry and Molecular Diagnostics. 4th ed. St. Louis, MO: Elsevier Saunders: 2006; chap. 26.Ridker PM, Rifai N, Rose L, et al. Comparison of C-reactive protein and low-density lipoprotein cholesterol levels in the prediction of first cardiovascular events. N Engl J Med. 2002;347:1557-1565.Sniderman AD. Differential response of cholesterol and particle measures of atherogenic lipoproteins to LDL-lowering therapy: Implications for clinical practice. J Clin Lipidol 2008;2:36-42.Tsimikas, S, Brilakis ES, Miller ER, et al. Oxidized phospholipids, Lp(a) lipoprotein, and coronary artery disease, N Engl J Med: 2005;353:46.Tsimikas S, Bergmark C, Beyer RW, et al. Temporal increases in plasma markers of oxidized low-density lipoprotein strongly reflect the presence of acute coronary syndromes. J Am Coll Cardiol. 2003; 41: 360.Tsimikas, S, Lau HK, Han KR, et al. Percutaneous coronary intervention results in acute increases in oxidized phospholipids and lipoprotein(a): Short-term and long-term immunologic responses to oxidized low-density lipoprotein. Circulation. 2004;109, 3164.Tsimikas S, Witztum JL, Miller ER, Sasiela WJ, et al. High-dose atorvastatin reduces total plasma levels of oxidized phospholipids and immune complexes present on apolipoprotein B-100 in patients with acute coronary syndromes in the MIRACL trial, Circulation: 2004;110, 1406. Walldius G, Jungner I, Holme I, et al. High apolipoprotein B, low apolipoprotein A-I, and improvement in the prediction of fatal myocardial infarction (AMORIS study): a prospective study. Lancet. 2001;358:2026-2033.Yusuf S, Hawken S, Ounpuu S, et al. Effect of potentially modifiable risk factors associated with myocardial infarction in 52 countries (the INTERHEART study): case-control study. Lancet. 2004;364:937-952.

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Nuclear Magnetic Resonance

The nuclear magnetic resonance (NMR) spectroscopy technique that was developed by LipoScience (LipoScience, Inc., Raleigh, NC), exploits specific magnetic properties of lipoproteins. This technology does not require separation of lipoproteins; serum or plasma can be run through the NMR sensor probe and all lipoproteins can be measured directly and homogeneously. The NMR platform works by subjecting the patient sample to a pulse of radio energy within a strong magnetic field. The energy that is given off by the lipids in the sample results is a signal that can be analyzed by the instrument to determine the number and size of lipoproteins present. Lipids associated with larger lipoproteins produce a signal that is distinct from those of smaller lipoproteins. A computer algorithm developed by LipoScience deconvolutes the signals into lipoprotein subclasses and then quantifies the number of particles in each class.NMR provides a useful and novel way to quantitate lipoprotein particles. However it is currently a proprietary technology and NMR analyzers are not yet readily-available for purchase and use in smaller clinical laboratories.

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Measuring particle number instead of cholesterol content has which of the following features or limitations?View Page

First Aid
Purpose of this Course

This program has provided some basic information about emergency situations that can occur in a laboratory.However, it is not a substitute for a hands-on first aid course, such as those that may be given at a local community center.The more you know about first aid, the more prepared you will be to act calmly and effectively in emergency situations, possibly saving lives.

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Fundamentals of Hemostasis
Which of the following laboratory tests of hemostatic function is a screening test used to assess the functionality of both the intrinsic and common pathways?View Page
What laboratory test result is commonly used to monitor oral anticoagulant therapy?View Page
Laboratory Tests of Hemostatic Function

Coagulation tests provide information that is used in diagnosing coagulation disorders, evaluating hemostatic function prior to surgery, and monitoring the effectiveness of anticoagulant therapy.

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Laboratory Tests of Hemostatic Function

Specimen rejection criteria established by your laboratory should be followed at all times, as improperly collected or processed coagulation specimens could adversely affect patient results. Generally speaking, hemolyzed specimens should not be used in coagulation testing because ADP liberated from lysed red blood cells can interfere with a number of coagulation tests, especially those involving platelet assessment. Grossly lipemic specimens may cause erroneous results or a clot may not be detected if a photo-optical coagulation system is used. An alternative method that is not affected by lipemia, such as an electromechanical method, may be required One way to avoid a grossly lipemic specimen is to ask the patient to fast prior to specimen collection.

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Laboratory Tests of Hemostatic Function – Prothrombin Time

The prothrombin time is a screening test that helps to assess the functionality of both the extrinsic and common pathways. The effectiveness and presence of factors I, II, V, VII, and X are assayed in this diagnostic test, as they are all found in the aforementioned pathways. The results of the prothrombin time are used in conjunction with other diagnostic tests, as well as the clinical picture of the patient, to determine any hemostatic abnormalities which may be present. In addition to being an integral part of the coagulation disorder assessment process, the PT is also used to determine therapeutic effectiveness of oral anticoagulants, by monitoring drugs such as Warfarin, Coumarin, and Dicoumarol. Prothrombin time test results are reported as the number of seconds needed for a clot to form in the patient specimen using the laboratory's instrument/reagent system, and as the International Normalized Ratio (INR).

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Laboratory Tests of Hemostatic Function – Prothrombin Time

The INR component of the laboratory result is a calculated value that is used by the clinician to monitor anticoagulant therapy and adjust dosage as dictated by clinical status. An INR of 2.0 - 3.0 is often desired as the therapeutic range. The following formula is used by the clinical laboratory to derive an INR value. The INR must be adjusted for every new lot of PT reagent. INR= (PT of patient/PT of geometric mean of the normal population)ISI The International Sensitivity Index, or ISI value, is provided by the reagent manufacturer as the relative sensitivity of the reagent itself. The INR is used to standardize PT results, and in turn, anticoagulant therapy, across laboratory instrumentation, methodologies, and locale. Be sure to frequently check that ISI values match those of the lot currently in use as erroneous results may otherwise occur .

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Laboratory Tests of Hemostatic Function - APTT

The activated partial thromboplastin time (APTT) is a screening test that helps to assess the functionality of both the intrinsic and common pathways. The effectiveness and presence of all the coagulation factors are assayed by this diagnostic test with the exception of factors VII and XIII. The results of the activated partial thromboplastin time are used in conjunction with other diagnostic tests, as well as the clinical picture of the patient, to determine hemostatic abnormalities which may be present. In addition to being an integral part of the coagulation disorder assessment process, the APTT is used to determine therapeutic effectiveness of heparin administration. Activated partial thromboplastin time results are presented to the clinician in seconds- the actual time elapsed until a clot was detected using the laboratory's instrument/reagent system.

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Fundamentals of Molecular Diagnostics
Overview

Molecular diagnostics have begun to play an integral part in clinical laboratory diagnostic testing. Traditionally, molecular diagnostics have been utilized in three major clinical areas: Infectious diseases Genetics Tumor markers These molecular based diagnostic tests, while historically reserved for specialty/reference labs, have recently seen expansion of their utility within the scope of routine clinical laboratories. Molecular based diagnostics can be utilized by small labs as well as large ones, and can be found in virtually every department of the clinical laboratory.

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Resources

It is imperative to follow the individual package insert procedures when collecting and handling specimens. Reference labs provide specimen requirements as well as collection, handling and transport guidelines.The Clinical and Laboratory Standards Institute (CLSI) formerly known as NCCLS: National Committee for Clinical Laboratory Standards has published procedures for collection including those specific to molecular diagnostics.

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Transport

Many clinical laboratories utilize reference laboratories for molecular methodology testing. Transport and shipping of biological specimens must follow laws and regulations governing these types of specimens. Consult your laboratory’s accrediting agency and reference lab for specific polices and procedures.

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Selection

Selection of the specific methodology will be dependent on each individual laboratory’s needs assessment of various factors including: Turn around time Cost Patient population Personnel competency Laboratory equipment requirements Laboratory space requirements Test complexity Result reporting

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Infectious Diseases

Molecular methodologies can be useful in the detection of a variety of diseases that are important public health issues such as:Chlamydia trachomatis (CT) Neisseria gonorrhoeae (GC)Human papillomavirus (HPV)Human Immunodeficiency Virus (HIV)Herpes Simplex Virus (HSV)Cytomegalovirus (CMV)In many clinical laboratories, traditional methods have been replaced by molecular methodologies because testing can occur for several pathogens in a single specimen. This is termed multiplex testing.

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Disadvantages of Molecular Testing

Molecular methodologies while highly advantageous do contain limitations and certain disadvantages. These can include:Cost: Molecular methodologies are usually more expensive than standard traditional methodologies. Equipment and reagent costs could be prohibitive to some laboratories. As molecular methods become more standard, the costs could potentially decrease. Currently, laboratories that consider the cost prohibitive prefer to transport molecular specimens to a reference laboratory.Personnel requirements: Depending on laboratory accreditation requirements and testing methodologies some personnel may not be qualified to competently perform molecular testing. Laboratory space requirements: Molecular amplification methods require dedicated space that may not be available in some clinical laboratories.

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Which of the following are considered advantages of molecular testing?View Page
References

Burtis CA, Ashwood ER, Bruns DE, eds. Tietz Textbook of Clinical Chemistry and Molecular Diagnostics. 4th ed. St. Louis, MO: Elsevier Inc; 2006.Clinical and Laboratory Standards Institute (CLSI). Collection, Transport, Preparation, and Storage of Specimens for Molecular Methods; Approved Guideline. CLSI document MM13-A. NCCLS. Wayne, PA: 2005.Clinical and Laboratory Standards Institute (CLSI). Molecular Diagnostic Methods for Infectious Diseases; Approved Guideline. Second ed. CLSI document MM3-A2. NCCLS. Wayne, PA: 2006.

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Hereditary Hemochromatosis
Introduction

Hereditary hemochromatosis (HH) is a disorder of iron regulation that results in excessive dietary iron absorption through the gastrointestinal tract. Over time, the resultant iron overload and its deposition in tissue may lead to widespread organ damage, a variety of chronic disorders, and even death. Although it is a genetic disorder, clinical symptoms most typically become apparent in middle aged adults. Iron overload occurs in a variety of hereditary and acquired forms, known as iron storage diseases. HH is the most common cause of inherited iron overload. (1) Due to lack of awareness, HH often goes undetected or unrecognized by health care providers. Early detection to prevent the serious complications associated with iron overload has important consequences for reducing morbidity and mortality. Laboratory tests that assess iron levels and molecular assays for genetic mutatations are essential for both its detection and diagnosis.

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General Overview of Testing

Tests for evaluating iron metabolism are generally used as initial or screening tests for hereditary hemochromatosis (HH) as they will detect the phenotypic expression of HH. These tests include serum iron (SI), transferrin (Tf) or total iron binding capacity (TIBC), serum ferritin (SF), and unsaturated iron binding capacity (UIBC).The serum ferritin assay is also used to assess the effectiveness of HH treatment.Molecular (DNA) analyses for HFE mutations are considered to be confirmatory tests for HH which may be ordered reflexively in patients with elevated iron results. Laboratories should establish their own reference intervals for assays of iron metabolism. In general, reference intervals vary by sex and by method used for the assays discussed in the following section. Typical reference intervals are included in the following sections for instructive purposes only and should not be used for evaluating actual patient data.The results of laboratory tests assessing iron metabolism should be interpreted with caution because a number of pre-analytical and physiologic factors can affect the results. Repeating elevated test results on fasting specimens is often advisable.

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Which laboratory assay is considered to be a confirmatory test for hereditary hemochromatosis (HH)?View Page
Specific HFE Mutations

Several mutations of the HFE gene have been described. In the C282Y mutation, a base substitution leads to a change in the amino acid in position 282 from cysteine (C) to tyrosine (Y). The loss of the sulfhydryl-containing amino acid disrupts the tertiary structure of HFE so that it no longer binds to beta-2 microglobulin. Beta-2 microglobulin appears to act along with other proteins to chaperone the newly synthesized HFE out of the Golgi apparatus and to the cell surface where it can then bind to TfR. In the C282Y mutation, HFE remains in the Golgi, never making it to the cell surface. The result is that transferrin binding to TfR is enhanced and excessive amounts of iron enter the cells of the small intestine, liver, and other tissues. A second mutation, H63D, causes a histidine (H) residue in position 63 to be replaced by aspartic acid (D). The mechanism by which this mutation leads to increased iron uptake is less well understood when compared to the C282Y mutation. Unlike the C282Y mutation, the H63D mutation does not seem to affect the binding of beta-2 microglobulin and intracellular movement, since detectable concentrations of the mutated protein are found on cell membranes. Some researchers speculate that the H63D mutation affects the binding of proteins involved in iron regulation and uptake at the cell surface.A third mutation, S65C, leads to a serine-to-cysteine substitution in its associated protein. This mutation has been been found in some compound heterozygotes for C282Y or H63D, but is rarely associated with iron overload in HH.Additional mutations of HFE have been identified, but their clinical significance is unclear. Most laboratories performing molecular assays test for only the C282Y, H63D, and S65C mutations.

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General Clinical Considerations

Hereditary hemochromatosis (HH) is frequently discovered only during management of associated illness or routine health evaluations. It has been estimated that only a small percentage of all affected persons are actually diagnosed. Individuals with HH may be symptomatic for several years prior to diagnosis and may have consulted multiple health care providers.Under-diagnosis of HH is thought to occur due to:• Lack of specificity of early signs and symptoms• Asymptomatic status of some patients until damage to organs and tissues has occurred• Confusion with liver disease due to other causes• Insufficient awareness and knowledge of HHEarly identification of persons with HH is essential to prevent serious and irreversible complications associated with severe iron overload. A classic triad of skin hyperpigmentation (bronzing), type 2 diabetes, and hepatic cirrhosis has long been recognized as evidence of advanced iron overload. However, persons with HH may present with a much wider variety of signs and symptoms, particularly if they are seen before significant iron accumulation has occurred. Age of presentation and disease severity are highly variable. A diagnosis of HH is based on laboratory evidence of iron overload, genetic mutations associated with HH, and presence of clinical signs and symptoms consistent with HH.(10)

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Secondary Disorders of Iron Overload

In addition to hereditary hemochromatosis (HH), there are other conditions of iron overload that must be considered in a differential diagnosis. Disorders such as sickle cell disease, thalassemia, sideroblastic anemia, congenital dyserythropoietic anemia, and liver disease may also cause iron overload. Transfusion-dependant patients and persons who abuse iron-containing vitamin supplements are also at risk. These conditions are usually described as secondary iron overload, in contrast to the primary iron overload of HH.Patient history, clinical signs and symptoms, biochemical and hematologic laboratory analyses, and possibly results of a liver biopsy may be needed to establish a diagnosis of a condition causing secondary iron overload. DNA tests for common HFE mutations are very likely the most important diagnostic tool for identifying HH as the cause of iron overload. In some patients, both secondary causes and HH may be contributing to iron overload. Differentiating the secondary causes of iron overload from HH is heavily dependent on the results of laboratory assays, but a complete discussion is beyond the scope of this course.

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Diagnosing HH

The diagnosis of hereditary hemochromatosis (HH) is made through a combination of laboratory tests and medical evaluation of a patient's signs and symptoms. Iron overload is identified by tests that evaluate iron metabolism, while molecular assays are needed to document mutations in the HFE gene or others such as hepcidin, hemojuvelin, or transferrin receptor. Individuals with documented iron overload who exhibit signs and symptoms consistent with HH and who possess HFE or other mutations are considered to have HH. Other causes of secondary iron overload may need to be ruled out.An example of a testing algorithm is shown.

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Which of the following is (are) needed for a diagnosis of hereditary hemochromatosis (HH)?View Page
What laboratory test reflects circulating iron that is bound to transferrin?View Page
Transferrin Saturation

Transferrin saturation (TS) is usually reported along with the SI and TIBC. TS indicates the percent of iron binding sites on transferrin that are carrying iron. TS is derived from a calculation using the formula:TS =(SI/TIBC) x 100TS results are reported as percentages. Typical reference intervals for TS are 20% to 55% for males and 15% to 50% for females. TS is generally considered to be the most sensitive laboratory test for detecting altered iron metabolism in hereditary hemochromatosis (HH). It may be elevated prior to significant deposition of tissue iron. TS levels increase as additional iron is accumulated.A drawback to using the TS is that it is dependent on performing both the SI and TIBC. The UIBC (see section below) may be a lower cost alternative.The optimal TS criterion for detecting HH is controversial. Using a TS of >60% for males and >50% for females has been found highly accurate in detecting abnormal iron metabolism in persons with HH. Others studies suggest using lower TS levels, e.g. 45%, as a criterion indicating further testing is warranted. Current guidelines from the American College of Physicians include a TS cutoff level of >55% for identifying iron overload. (11)Patients with initially increased TS should be followed by performing a second TS from a fasting morning specimen. The patient should also be advised not to take vitamins supplemented with iron or oral contraceptives for several days prior to the repeated test. TS levels may be affected by diurnal variation, dietary factors, and co-existing disease states such as inflammation and hepatitis. Patients with HH may have falsely normal TS if chronic blood loss or inflammatory disease is present.

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Why is serum ferritin (SF) a less than optimal screening test for hereditary hemochromatosis (HH)?View Page
Screening Controversies

The subject of screening for hereditary hemochromatosis (HH) is controversial and is currently being debated in the medical literature. Using laboratory tests to screen the asymptomatic general population is currently not recommended due to issues of testing costs, low genetic penetrance, and the possible risk of discrimination. Targeted case finding in select high risk populations such as men of Northern European ancestry may be a better approach to screening. (12)Molecular-based (DNA) assays required for confirmation of HH are costly when used for general population screening. Because recent studies have shown that a high percentage of persons with C282Y mutations do not develop iron overload or HH-related clinical conditions, screening for these mutations may falsely label an individual with a disease diagnosis. At the present time, it is impossible to determine which homozygotes or heterozygotes for HFE mutations will eventually develop iron overload. Furthermore, there is potential risk of discrimination in obtaining health insurance for persons identified as having genetic disorders.In contrast, some experts do advocate for screening the general population. Mutations associated with HH are very common in Caucasians in the US. Individuals who know they carry mutations associated with HH may benefit from periodic testing for iron overload. Finally, laboratory tests that assess iron status are relatively inexpensive, widely available, and offer one approach to screening for phenotypic expression of HH. Screening first-degree family members of a person with documented HH is generally considered to be worthwhile. Early detection of HH in relatives with common mutations may permit treatment before the development of substantial iron overload and related disease due to organ damage.

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What is one established reason supporting general population screening for hereditary hemochromatosis (HH)?View Page
UIBC

Unsaturated iron binding capacity (UIBC) may also be used as a marker for altered iron metabolism. UIBC represents the portion of iron binding sites on transferrin that are not occupied by iron. Therefore, a low UIBC indicates that transferrin is highly saturated with iron, a finding consistent with hereditary hemochromatosis (HH). HH may be suspected when the UIBC is less than 143 micrograms/dL, a criterion suggested by the results of one study.(5)UIBC may be a lower cost alternative to the more complex transferrin saturation (TS). UIBC and SI are both fully automated procedures that are available on widely used laboratory instruments. The TIBC can be calculated by adding UIBC and SI, resulting in a value for TIBC that can be used for determining TS: TS = SI/(SI + UIBC) X 100

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Molecular Tests

DNA tests for HFE mutations associated with hereditary hemochromatosis (HH) are available in some clinical laboratories and reference laboratories. Testing for the presence of the C282Y is essential, although most labs also test for H63D and S65C mutations. Molecular testing is most appropriate for confirmatory testing of symptomatic individuals with altered iron studies (increased TS and SF), in pre-symptomatic individuals (increased TS, normal SF and liver function tests), and in family members of individuals diagnosed with HH. The use of genetic tests alone for routine screening of asymptomatic persons is not recommended for several reasons. A positive test indicating the presence of HFE mutations does not guarantee that an individual will develop clinically significant iron overload or predict severity of symptoms. A negative result (no HFE mutations present) does not rule out a diagnosis of iron overload because of genetic heterogeneity. Compared to biochemical analyses for iron, molecular assays are expensive. Finally, molecular testing may result in the diagnosis of a genetic disease, thus opening up the possibility for discrimination in health insurance coverage. Using molecular methods, DNA is extracted from leukocytes in whole blood samples or from buccal cells and analyzed for specific HFE mutations using polymerase chain reaction (PCR) with melt curve analysis. Currently there are no FDA-cleared products for HFE testing, and testing laboratories are using "home brew" reagents. This situation is expected to change as manufacturers submit products for FDA approval.

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HIPAA Privacy and Security Regulations
Case Study: Administrative Safeguards You are the technologist in charge of the hematology section in a hospital laboratory and you are reviewing blood count results for 100 patients as part of an internal quality assurance project. You review the clinical findings in the electronic medical record to correlate with the laboratory results. The following week, you get a call from your hospital security officer. She says that a routine computer system audit has revealed that you accessed the records of 100 patients and she would like to know why.You tell her:View Page
Minimum Necessary Use and Disclosure

Minimum Necessary means that the laboratory will use and disclose only the minimum PHI necessary to accomplish its intended purpose, such as resulting the requested test. The regulation recognizes that there are situations where all of the PHI on a patient can be released. These include: When releasing PHI to another covered entity for treatment. When releasing an individual's PHI to himself or herself. When an individual has signed an authorization to release the PHI. When required to do so by law.

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Authorization

The privacy regulations give covered entities permission to use and disclose PHI for treatment, payment, and health care operations (TPO), without obtaining specific authorization.A covered entity may disclose PHI to other covered entities such as reference laboratories, and homecare services, which are providing services to the primary covered entity.The service that the other covered entity is providing must fall within treatment, payment or health care operations (TPO).If the service being provided does not fall within TPO, an authorization is generally required.An authorization form must state the specific disclosures of PHI to be made, what the information will be used for, and must be signed and dated by the patient.

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Case Study: Authorization You are working in a physicians office. The doctor orders laboratory and other diagnostic tests on a patient with suspected Alzheimer's disease. He then asks you to give the patient's name and contact information to the local Alzheimer support group without getting permission from the patient or his legal guardian. Does the doctor need authorization from the patient or his legal guardian to do this?View Page
Case Study: De-identified Health Information. You work in a laboratory microbiology department which provides a local nursing home with information about the effectiveness of various antibiotics it uses to treat infections. You print the requested information, including complete patient identification, bacterial organisms identified, and their sensitivity to various antibiotics. What information should you provide to the nursing home?View Page
Case Study: Limiting Use & Disclosure of PHI You are the customer service representative in a clinical laboratory. You get a call from someone at a local gastroenterologist's office, with whom you are personally familiar, requesting that you fax results on a patient, which the referring physician's office had failed to provide. The doctor needs the test results immediately. Under the HIPAA Privacy Regulations the you can comply with this request, without getting written authorization from the patient.View Page
Case Study: Incidental disclosures and safeguards. As a manager, you guided a group of high school students through your clinical laboratory during a field trip. You did not explain the laboratory's privacy policy to the teacher and students, because you thought they would have little access to PHI. However during the tour, the students overheard names of patients and blood tests, saw laboratory reports laying on desks, and viewed test results on computer screens. This is acceptable under the HIPAA Privacy Regulation since these were incidental disclosures that could not reasonably be prevented.View Page
Case Study: Minimum Necessary Use and Disclosure You are a ward clerk responsible for inserting laboratory reports into patients' medical records (charts). You open the chart directly to the laboratory tab, insert the report, and avoid "paging through" the entire medical record. "Paging through" and browsing the medical record to satisfy your curiosity would be a violation of the privacy regulations.View Page
Importance of Privacy - An Example

You will have many opportunities to avoid disclosing protected health information. Here is one simple example: Your best friend asks you to look up her mother's laboratory results. Knowing the HIPAA privacy regulation and your own departmental policies and procedures, you do not disclose the protected health information which she is requesting. You politely tell your friend that your are not allowed to give her the laboratory results.

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Who does HIPAA apply to?

HIPAA applies to: Health Plans (such as health insurance companies) Healthcare Clearinghouses (such as billing companies), and Healthcare Providers (including doctors, hospitals, laboratories, and pharmacies). HIPAA refers to these 3 groups as covered entities.

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HIV Safety for Florida
A person commits a misdemeanor of the first degree by:View Page
Reporting results

Each laboratory that performs a test indicative of HIV or AIDS shall report to the county health department in less than 2 weeks.To assure the confidentiality of the patient, reporting of HIV infection and AIDS must be conducted using a system developed by CDC or equivalent system.Each person who violates reporting rules may be fined $500 per offense.

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Overview

Prevention of HIV exposure is the best line of defense to prevent occupational transmission of HIV as there is no vaccine available to develop specific immunity and the postexposure prophylaxis is toxic. Following appropriate workplace practices in the laboratory focus on preventing needlesticks or other sharps injuries and exposure of mucous membranes and abraded skin to HIV-infected blood or body fluids.

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The following workplace practices minimize risk of HIV exposure to mucous membranes or abraded skin:View Page

Introduction to Bioterrorism
The Laboratory Response Network (LRN) is a multilevel system designed to link frontline clinical laboratories to advanced capacity testing laboratories. The frontline microbiology laboratories are classified by the LRN as:View Page
Members of the chemical component of the LRN define their network participation with a designation of level 1, 2, or 3. The level primarily responsible for working with hospitals and private laboratories is:View Page
Advantages of using Biological Agents as WMDs

They are easily available.Biological pathogens can be obtained from nature, hospital  laboratories, university research facilities, etc.They can be hard to detect.Small quantities can have potentially deadly or incapacitating effects on a susceptible population.They can be used covertly.They can be spread throughout large areas by natural convection, air or water currents. They can be easily spread.Ventilation systems in buildings is one way biological agents may be spread. In addition, transportation facilities could become part of the dissemination system by carrying biological agents far from their initial source.

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Laboratory Response

The broad base of clinical laboratories in this country is an essential component of our nation’s public health and healthcare system and is an essential link in addressing biological and chemical terrorism. In 1999 the Centers for Disease Control and Prevention (CDC) initiated the concept of a Laboratory Response Network (LRN).  The LRN is a network of local, state, federal, and military laboratories across the United States and internationally which work together in an integrated and coordinated way for a rapid response to public health emergencies. The LRN concept of operations is based on a system of safety and proficiency.

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The LRN Pyramid

The LRN is a multilevel system designed to link frontline clinical microbiology laboratories and hospitals and other institutions to state and local public health laboratories in supporting advanced capacity public health, military, veterinary, agricultural, water and food testing laboratories at the federal level. Laboratories within the LRN are divided into 3 levels: Sentinel Labs, Reference Labs, and National Labs.

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Reference Labs

Next up on the pyramid are the reference laboratories. These laboratories are sometimes referred to as “confirmatory reference” laboratories. They can perform tests to detect and confirm (rule-in) the presence of a threat agent. These labs ensure a timely local response in the event of a terrorist incident. Rather than having to rely on confirmation from laboratories at CDC, reference laboratories are capable of producing conclusive results. This allows local authorities to respond quickly to emergencies.

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National Labs

At the highest level are the “national” laboratories. Examples would include those operated by CDC, the United States Army Medical Research Institute for Infectious Diseases, and the Naval Medical Research Center. These laboratories have very unique resources to handle highly infectious agents and the ability to identify specific agent strains.

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Laboratory Response - Chemical

Currently there are over 60 territorial and metropolitan public health laboratories that are members of the chemical component of the LRN. A designation of Level 1, 2, or 3 defines their network participation.

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Laboratory Response - Chemical, Level 3

Level 3 laboratories are responsible for: Working with hospitals and private laboratories in their jurisdiction Knowing how to properly collect and ship clinical specimens Ensuring that specimens, which can be used as evidence in a criminal investigation, are properly handled and that chain-of-custody procedures are followed Being familiar with chemical agents and how they can affect health and well-being Training on anticipated clinical sample flow and shipping regulations Working to develop a coordinated response plan for their respective state and jurisdiction

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Laboratory Response - Chemical, Level 2

In addition to the responsibilities listed for Level 3, over 40 laboratories also participate in Level 2 activities. At this level, laboratory personnel are trained to detect exposure to a limited number of toxic chemical agents in human blood or urine, the analysis of cyanide and toxic metals in human samples, for example.

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Laboratory Response - Chemical, Level 1

At present, 5 laboratories participate in Level 1 activities. At this level, technical personnel are trained to detect exposure to an expanded number of chemicals in human blood and urine. This includes all Level 3 and 2 laboratory analyses, plus analyses for mustard agents, nerve agents, and other toxic chemicals.

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In the LRN hierarchy, large organizations like the CDC, the United States Army Medical Research Institute for Infectious Diseases, and the Naval Medical Research Center are classified as View Page
In the Chemical Component of the LRN, there are 60 laboratories divided into 3 levels. At the top, with unique facilities unavailable to the level 1 - 3 labs, is/are theView Page
Sentinel Labs

The frontline clinical microbiology laboratories are known as “sentinel laboratories”. The sentinel laboratories play a key role in the nation’s preparedness efforts. These laboratories perform the initial screening of clinical specimens for potential pathogens (rule-out) and refer specimens or isolates to a state or local public health laboratory at the reference level of the LRN. There are two kinds of sentinel laboratories: advanced and basic. Classification depends on their biological safety level and analytical capability.

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Your Response – At Work

Recent events, including the terrorist attacks on September 11, 2001 and the subsequent bioterrorist releases of anthrax, have been a harsh awakening that the nation’s workplaces could be terrorist targets.Traditionally laboratory safety guidelines have emphasized use of optimal work practices, appropriate containment equipment, well-designed facilities, and administrative controls to minimize risks of unintentional infection or injury for laboratory workers. Today, in addition to the above, laboratories must make a risk and threat assessment, secure data and electronic technology systems, plus develop policies regarding specimen accountability, facility security, and emergency response.The next few pages will cover a number of things that you can do to assist in making your laboratory more risk free to a terrorist attack and some things you can do in case that security is breached. You too have a role in the security of your workplace!

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Introduction to Bone Marrow
Preparation of Concentrated Smears

In some laboratories the anticoagulated sample is used to prepare concentrated smears. Placing the fluid in a Wintrobe tube and centrifuging it separates the sample into four layers:fat and perivascular cellsplasmabuffy layer - myeloid and nucleated erythroid cellserythrocytesThe volume of each layer is measured using the scale on the Wintrobe tube and then the percentage of each layer is calculated. Next the plasma is removed and a smear is made from the buffy coat and top of the red cell layer. Either the manual push method or cytospin technique may be used to make the smears. They may be stained with a variety of cytochemical stains. Concentrated smears are used to examine cell morphology and demonstrate the presence of abnormal cells when the marrow is hypocellular. The smears cannot be used for differential counts or evaluation of cellularity.

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Introduction to Quality Control
Which of the following types of controls can be used to measure accuracy of a laboratory's methods?View Page
Quality Control Procedures

Quality control procedures may include the following: monitoring variables within the laboratory, like water quality, glassware calibration, and instrument calibration, that may affect results defining protocols for performing quality control for each test procedure participating in proficiency testing programs graphing quality control results performing daily and periodic analysis of quality control data and charts troubleshooting quality control errors documenting quality control errors, including the steps taken to resolve the problem and verification of success performing routine maintenance of laboratory instruments keeping records

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Federal Regulations

The importance of quality control is recognized by the federal government. Federal regulations, such as the Clinical Laboratory Improvement Amendments of 1988 (CLIA 88), require successful participation in external quality control programs for clinical laboratories, regardless of size. These regulations are aimed at providing all citizens with the highest quality of health care and at the same time controlling costs.

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Internal Quality Control

Internal quality control is set up within a laboratory to monitor and ensure the reliability of test results from that laboratory.The primary tool for internal quality control is called a control. A control is a specimen with a predetermined range of result values, called control values, that is processed in the same manner as a patient sample. Control samples are processed with each series or run of patient samples.If the result of a test on a control sample is different from its known value, this indicates a problem in the equipment or the methods being used.

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Assayed and Unassayed Controls

Commercially prepared controls come in either assayed or unassayed forms. Assayed controls are tested by multiple methods before sale, and are sold with the results of the tests. Assayed controls: are more expensive than unassayed controls are used to evaluate accuracy and precision avoid laboratory errors in determining control values may only be suitable for specific methods or conditionsWhile the manufacturer's control values can be used to some extent to measure accuracy, the best measure of accuracy is certified reference material.Unassayed controls are not tested by the manufacturer before they are sold. The control values for these materials must be determined by the individual laboratory. Unassayed controls: are less expensive than assayed controls are used to evaluate precision only avoid manufacturer error in determining control values control values are customized to the laboratory's own methods and conditionsA final note: although commercially available control materials are screened for hepatitis antigens and HIV antibodies, control materials should still be handled with precautions, since they contain biological materials and could contain infectious agents.

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Additional Variables

To ensure reliability of results, many other factors besides controls are monitored. These include: water quality analytic balance calibration glassware calibration centrifuge calibration thermometer calibration electric power stability heating bath temperatures refrigerator temperatures freezer temperatures expiration of reagents, standards, and controls instrument maintenance procedure manuals, including written step-by-step procedures of all tests performed by laboratory method selection, based on local population and clinician needs normal range verification based on the local population technical competencies of laboratory staff members

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Instrument Maintenance

Instruments require regular maintenance by laboratory personnel, and sometimes by qualified repair technicians. The instrument manufacturer will provide recommendations for how often to perform maintenance; those recommendations should be followed. Generally, a log book will document the details of the maintenance: date time name of employee or service engineer description of maintenance done, including listing any parts replaced description of problems encountered description of steps taken to resolve the problem and verify the instrument is functioning, after the changes were madeInstrument logs should be regularly reviewed by supervisors to monitor instrument performance. By reviewing the logs, the supervisor can make changes to the maintenance schedule, based on the frequency of breakdowns.

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External Quality Control

External quality control is performed to ensure the reliability of test results between different laboratories. It is also required by CLIA for laboratory accreditation. External quality control is generally accomplished through proficiency testing (PT). In proficiency testing, simulated patient samples are sent out to laboratories for testing. The CLIA standards for handling proficiency testing specimens are as follows: PT samples must be tested with the laboratory's regular patient load. PT samples must be tested the same number of times that patients' samples are tested routinely. Laboratories participating in PT programs must not engage in interlaboratory comparison of PT sample results. Laboratories may not send PT samples to another laboratory for analysis. Laboratories must document all steps of processing for PT samples. PT is required for only the primary method used for testing of analytes in patients' samples during the period covered by the PT event.In return for their participation, the laboratory will receive the following information: results for each analyte sample mean result for each analyte standard deviation of results by the comparative method number of laboratories using the same method standard deviation index (SDI) lower and upper limits of acceptability of resultsPT results that are between the lower and upper limits of acceptability are considered satisfactory. For chemistry, 80% of samples must test within the acceptable range for the PT to be considered successful. External quality control serves several purposes, including: providing a check on internal quality control detecting errors in a lab's methods providing a comparison of testing methods, which is useful in selecting new methods

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Which of the following statements about external quality control are true?View Page

Introduction to the ABO Blood Group System
In what way are the ABO serum antibodies unique among blood group systems?View Page

Laboratory Ergonomics
Creating an Ergonomically Safe Work Environment

Both the employer and the employee should share the responsibility for assessing and improving ergonomics in the laboratory. A three-step ergonomic program includes: Finding the hazard Determining what improvements / changes should be made Taking action to improve the workplaceEmployers should: Provide ergonomics education Provide ergonomically designed tools and equipment Allow frequent stretch breaks If possible, adjust work schedules to prevent employees from performing repetitious tasks for prolonged periods of time. As an employee, you should evaluate the ergonomic practices in your work area.Employees should: Understand the risk of injury Apply ergonomic principles to the performance of tasks Look for ergonomic hazards and improve the workplace whenever possible Recognize and report early signs of musculoskeletal disorders (MSDs)

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Scenario #3Jim spends most of his workday sitting on a stool at the technical workbench. The image on this page illustrates how he routinely sits. Lately, he has been experiencing lower back and leg discomfort that continues to bother him when he leaves work. He has been having trouble sleeping because of the pain in his legs. Eventually, the pain progresses to the point where he cannot work an entire day. What may have caused the problem and what could have been done to prevent the MSD from developing? Consider what could be the problem based on your observation of Jim's normal sitting position. Then click on the blue text below to see the ergonomic evaluation and possible solutions.View Page
Examine the image on the right. What problems can you see that may result in MSDs for this laboratory worker if her job involves frequent use of the computer for prolonged periods of time?View Page
Learn to Use Your "Ergonomic Eye"

Ergonomics is an important part of the overall laboratory safety program that ensures the well-being of all employees. The potential for MSDs while performing routine tasks exist in all areas of the laboratory. Laboratory workers should apply ergonomic principles to the performance of their job tasks and should report workplace design concerns. Each physical activity should be observed for opportunities to decrease physical stress and increase comfort.

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References

Centers for Disease Control and Prevention. Laboratory ergonomics. Available at: http://www.cdc.gov/od/ohs/Ergonomics/labergo.htm. Accessed July 6, 2009.Cornell University. CUErgo. Available at: http://ergo.human.cornell.edu/ Accessed July 6, 2009.National Institute for Occupational Safety and Health. Ergonomics and musculoskeletal disorders. Available at: http://www.cdc.gov/niosh/topics/ergonomics/ Accessed July 6, 2009.UCLA Ergonomics. Musculoskeletal disorders: Anatomy of an injury. Available at: http://ergonomics.ucla.edu/MSD_Anatomy.html. Accessed July 6, 2009.US Department of Labor. Healthcare wide hazards module: Ergonomics. Available at: http://www.osha.gov/SLTC/etools/hospital/hazards/ergo/ergo.html Accessed July 6, 2009.

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Repetitive Motion Injuries

Repetitive motions can cause a variety of disorders that affect nerves, tendons, and muscles. Symptoms can include tingling or numbness in the fingers or hands, decreased range of motion, decreased grip strength, sleep interupted by numbness or discomfort in the hands, pain in fingers, hands, or wrist, or pain shooting up into the forearms or arms.Some common afflictions that could affect laboratory workers due to the nature of their jobs are listed in the table below. Condition Symptoms Cause Carpal tunnel syndrome Pain that radiates up the arm, numbness or tingling in the thumb, index, or middle finger and weakness in the wrist and hand Compression of the median nerve that runs from the forearm into the hand Thoracic outlet syndrome Numbness and tingling in the hand, intensified with overhead activities Compression of the nerves and blood vessels between the neck and shoulder Radial tunnel syndrome Elbow pain, pain near the base of thumb, or pain anywhere in between. A common symptom is wrist weakness. Compression or entrapment of the radial nerve; may be caused by repetitive wrist and finger extension or repetitive forearm turning. Tendinitis Stiffness, tightness, and burning sensation; may experience a deep nonspecific pain. Grip impairment. Occurs most often in the tendons of the fingers, thumb, forearm, elbow, and shoulder. Repetitive motions or maintaining an awkward position that stresses tendons beyond their strength. Friction from overuse can cause inflammation. Tenosynovitis Pain, swelling, difficulty moving the joint in the affected area Inflammation of the tendon sheath

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Causes of Work-related Musculoskeletal Disorders

The primary goal of ergonomics is the prevention of musculoskeletal disorders (MSDs). There are many potential causes of MSDs. Injury can occur from a single event (strain, sprain, slip, or fall) or result from build-up of tissue damage from many small injuries. An MSD can develop over time if a motion is repeated consistently so that the constant trauma causes damage to a muscle, tendon, bone, or bursa of a joint. Force, vibration, or maintenance of an awkward position for a prolonged period of time can cause MSDs.Some specific causes of MSDs that are related to laboratory tasks are: Prolonged use of a keyboard or mouse Prolonged sitting at a microscope Pipetting Screwing and unscrewing vial caps Standing at a laboratory instrument for a prolonged period of time Lack of rest - intensive hours at the workstation with few breaks Sustained awkward position

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Laws and Rules of the Florida Board of Clinical Laboratory Personnel
Your Role

As a clinical laboratory worker, your role is vital in the health care process. You provide information to doctors, nurses, and healthcare organizations that is vital to proper patient care. Because your role is so important, you must be properly qualified, trained, and licensed for your position. You must also keep up with the latest laboratory techniques and developments by fulfilling continuing education requirements; and you are bound by a code of ethics, which ensures that patient results are accurate, reliable, and free from error and bias.

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Public health laboratory scientists

Public health laboratory scientists are also regulated by the Board. The table below outlines the various requirements for applicants to receive licensure for a public health laboratory. Public Health Laboratory RequirementsDirectorFulfill the same requirements as a clinical laboratory directorSupervisorBe certified by National Registry in Clinical Chemistry or American Society for MicrobiologyBe licensed as a technologistHave five year's relevant experiencePass the state examTechnician (microbiology)Have a Bachelor's degree in one of the biological sciencesObtain American Society for Microbiology or the National Registry in Microbiology Certification in Public Health Microbiology Technician (chemistry)Have a Bachelor's degree in one of the chemical, biological, or physical sciencesObtain National Registry of Clinical Chemistry Certification in Public Health ChemistryTechnician (conditional)Have a Bachelor's degree in one of the chemical or biological sciencesPerform tests only under the direct supervision of a licensed pathologist, director, supervisor, or technologist.Receives a conditional two-year license, which may be renewed only once A license from the Board of Clinical Laboratory Personnel allows you to work in a public health laboratory at the same level and specialty.

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Clinical laboratory personnel license

To practice as a clinical laboratory scientist in the state of Florida, you must have an appropriate Florida license. Without a license, you cannot conduct clinical laboratory examinations or report test results. You do not need a Florida license to work in: Laboratories run by the federal government. Labs that perform only waived testing. Labs run exclusively for research and teaching purposes that do not report patient results.These laboratories may have other licensing and training requirements.

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Technologist Qualifications

Meets one of the following:Bachelor's degree in clinical laboratory, chemical or biological science plus:Completion of a medical technologist training program ORThree years of laboratory experience, at least one of which must be in the applied-for specialtyAssociate's degree plus:Florida technician's license and completion of a technician level medical laboratory training program ORFive years of laboratory experience, at least one of which must be in the applied-for specialtyPasses an examination in one or more specialtiesCompletes one hour of HIV / AIDS continuing educationCompletes two hours of medical errors continuing education

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Director Qualifications

Effective date March 17, 2008All applicants for a Director license must meet the qualifications for a high complexity laboratory director that are defined in 42 CFR 493.1443 as published on October 1, 2007.A licensed physician may direct a clinical laboratory without a separate laboratory director's license if he / she is certified in clinical pathology by the American Board of Pathology (ABP) or the American Osteopathic Board of Pathology (AOBP); is board-certified in the pertinent laboratory speciality; and/or has four years of pertinent clinical laboratory experience (post-graduate) with two years experience in the speciality that will he/she will direct.A non-physician may obtain a director's license for a specialty area if he / she: Holds an earned doctoral degree in a chemical, biological, or clinical laboratory science Is certified in the pertinent laboratory specialty by an approved national board A director can oversee up to five laboratories.

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Director Responsibilities

A clinical laboratory director is responsible for the overall operation and administration of the clinical laboratory. The director can delegate responsibilities to licensed supervisors, but is ultimately responsible for the following: Ensuring the employment of personnel who have the necessary education and experience and who are competent to perform the procedures and tasks that are assigned to them. Overseeing performance and reporting of accurate test results Verifying the laboratory's compliance with federal and state laws, rules, and regulations Delegating certain administrative duties to supervisors Being available for on-site, telephone, or e-mail consultation Ensuring that test methods and procedures, quality control, and verification methods provide reliable and accurate results Ensuring compliance with quality control and quality assurance programs Ensuring enrollment and active participation of the laboratory in a proficiency testing program, monitoring proficiency testing results, and implementing corrective action when necessary Assessing laboratory staffing needs and advising management when insufficient clinical laboratory personnel are employed Selecting which tests the laboratory offers and which employees may perform them Establishing and maintaining a patient identification system for the laboratory Establishing and maintaining accurate billing practices

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Supervisor Responsibilities

A supervisor is the laboratory director's designee for monitoring compliance with all applicable regulations of the board and the department. Other duties include: Performing the duties of a technologist, if needed, in the specialty area(s) where licensure is held Assigning direct supervision responsibilities to licensed technologists if needed, while ensuring that direct supervision of technicians is properly performed. Evaluating technologists' and technicians' competency in running tests and reporting results Being available to all personnel to answer questions and resolve problems Providing day-to-day supervision of test performance, including on-site direct supervision of testing that is performed by technicians Ensuring that quality control is performed and corrective action taken if necessary Ensuring that no patient testing is reported until corrective action has been taken and the test system is properly functioning Providing orientation to all testing personnel Implementing a quality maintenance program

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Technologist Responsibilities

Technologists fulfill assigned supervisory responsibilities as needed and as authorized. Other duties include: Performing only those tests authorized by the director and for which the technologist is licensed by specialty. Following the laboratory's procedure for specimen handling, running tests, reporting results, and maintaining records Participating in proficiency testing and demonstrating that proficiency samples are tested in the same manner as patient samples Following quality control and instrument calibration policies Documenting corrective action taken when results exceed the laboratory's acceptable performance values Using professional judgment to ensure test validity, including recollecting and retesting samples that may be flawed or contaminated

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Technician Responsibilities

Technicians perform laboratory testing under direct and general supervision, as required by the test and the conditions of the technician's license. Other duties include:Performing tests only as authorized by the director and the technician's licensed specialty.Following the laboratory's procedure for specimen handling and running testsParticipating in proficiency testing and demonstrating that proficiency samples are tested in the same manner as patient samplesFollowing quality control and instrument calibration policiesDocumenting corrective action taken when results exceed the laboratory's acceptable performance valuesIdentifying potential problems with tests or report resultsNotifying a technologist or supervisor if results are outside the laboratory's acceptable performance levels

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Competency and Licensing Violations

Clinical laboratory personnel must be licensed and competent to perform their duties. This means holding the appropriate type of license for the task being performed (director, supervisor, technologist, or technician) and being certified in the appropriate specialty for any testing being performed. For example, an individual licensed as a technician in hematology may not perform the duties of a technologist in hematology, nor may that individual perform testing in the microbiology specialty. Showing a lack of competence to perform even licensed duties is a violation of Board rules. Consistent errors can tarnish a laboratory's reputation, and even a single error can harm patient care. Licensed personnel must be certain that they can perform their duties accurately and competently. All of the following are violations of Board rules:Performing clinical duties for which one does not hold a license.Performing services one knows one is not competent to perform.Showing lack of competence or making consistent errors in testing or reporting.Having a license revoked or suspended in another state.

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Kickback and Inducement Violations

Offering or taking a bribe, kickback, bonus, commission, or inducement is against the rules of the Board and against the law. Many companies give away small promotional items, such as pens or note pads, to promote their products. This is legal, but be cautious about accepting more valuable items. This could be seen as a bribe. All of the following are serious violations of Board, state, and federal rules:Participating in any commissions, bonuses, kickbacks, inducements, or split-fee arrangements from physicians, health care providers, suppliers, hospitals, nursing homes, other clinical laboratories, pharmacies, and other facilities.Exploiting or influencing a patient for financial gain, including promoting, selling, or withholding services, drugs, or referrals.

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Which of the following is NOT a responsibility of a clinical laboratory director?View Page
Which of the following is NOT a responsibility of a clinical laboratory supervisor?View Page
Which of the following are violations of Board rules?View Page
A director may only oversee one laboratory.View Page

Linear Regression Analysis

Medical Error Prevention
What does the JCAHO Speak UP campaign encourage?View Page
Which statement(s) are true?View Page
Which statement(s) describe potential causes of medical errors involving the blood bank?View Page
Which actions are sources of laboratory-related error?View Page
Which strategies help laboratory professionals prevent medical errors?View Page
The Joint Commission Sentinel Event Alert Since 1998, the Joint Commission has issued 25 Sentinel Event Alerts to the healthcare community. These publications include more than 50 evidence or expert-based recommendations for preventing adverse events. Sentinel Event Alerts address various error reduction topics: Transfusion reactions Inpatient suicide Infant abductions Wrong site surgery or other procedures Patient fallsLaboratory professionals can be involved in all of these types of Sentinel Events. The Joint Commission's first Sentinel Event Alert addressed the common practice of storing concentrated potassium chloride solutions in hospital nursing units. View Page
Direct Error Detection Even perfect systems designs cannot avert human limitations. Medical errors occur and they have to be detected before they can be resolved. Sometimes people directly observe and immediately report these mistakes.View Page
Awareness

Laboratory professionals can learn about medical errors and prevention by maintaining their awareness of laboratory-related news and events. They can read print and online news sources, magazines, and professional journals to stay informed. They can also learn what is happening through televised and other media reports. Staying up to date about current trends and progress in error prevention enables professional to learn from mistakes and prevent new ones.

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Study

The Internet provides extensive, current resources for studying ways to prevent errors. Performing word searches for Medical Errors or Patient Safety or Laboratory Errors identifies a wealth of Internet resources. A Patient Safety search lists more than 93 million items.

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Discussion

Laboratory discussion meetings help to prevent medical errors. The staff can meet periodically to discuss recent averted adverse events and ones that might have been averted.Discussion should not be about blame. Privacy must be protected, so real names should not be identified. Management can provide guidelines for discussion and analysis.A suggested format for discussion:1. Briefly describe each adverse event.2. Identify its possible causes.3. Discuss relevant guidelines.4. Suggest possible preventive actions.Discussion can include actions that do and do not work to prevent medical errors.

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These statements describe ways laboratory professionals can prevent medical errors.View Page
Joint Commission Patient Safety Goals Joint Commission adopted national patient safety goals for healthcare organizations, including specific goals for laboratories. 2009 Laboratory Services National Patient Safety Goals These goals are directly quoted.View Page
American Society for Clinical Laboratory ScienceThe American Society for Clinical Laboratory Science, ASCLS, joins the leadership effort to prevent medical errors and increase patient safety.View Page
Types of Medical Errors Medical errors usually belong to one or more of these categories:View Page
Where Errors Occur

Insurance industry analysis of data from 1985 through 2003 shows that about two thirds of medical errors occur in hospitals and about one third occur in physician offices. About half of hospital errors occur in operating rooms and one sixth occur in patient rooms. Wherever medical errors occur, laboratory professionals can be involved.

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Preanalytic Medical Errors

Medical errors are possible at any phase of patient care. Preanalytic medical errors begin with the patient and the places he or she receives medical care--the bedside, chair-side, hospital, clinic-- wherever the patient is located. The possibility for these errors continues through the ordering processes for medical tests or procedures. Preanalytic medical errors also happen with the systems, processes, and procedures involved in the collection of test samples from patients. These medical errors occur during the time before the laboratory is directly involved in assaying and analyzing test samples. Examples of preanalytic medical errors: Wrong patient Wrong test Wrong timing Wrong collection procedure Wrong tube, container, additive

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Analytic Medical Errors

Medical errors also occur in the analytic processes and systems of patient care. Analytic errors begin with problems in the transportation of medical samples for testing. These occur between the patient's location and the testing facility. They happen during the time between specimen collection and arrival in the testing facility. The possibility for analytic medical error continues through the analytic processes and procedures of medical testing. Analytic medical error also includes systems, processes, and procedures involved in the transmission and reporting of test results. These medical errors occur during the time the laboratory is directly involved in receiving, analyzing, and reporting test samples. Examples: Wrong transport storage or temperature Delay in transport Sample mixup during transport Acceptance of unacceptable samples that are insufficient, hemolyzed, or clotted Centrifugation, mixing, and other test sample preparation errors Wrong test procedures Test control errors Sample mixup during testing Outdated reagents Wrong reagents Test result mixup Transcription errors Data reporting process errors Result report delays

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Postanalytic Medical Errors

Errors also occur after analyses are completed and reported. Postanalytic errors begin with the medical professionals who receive test results, and they include interpretation of the results. These errors can occur at--the bedside, chair-side, hospital, clinic-- wherever the patient and the medical professional are located. The possibility for postanalytic medical error continues through diagnosis and treatment procedures and processes. These medical errors occur during the time after the laboratory reports test results. Examples: Wrong test value associated with patient Wrong test interpretation Wrong diagnosis Wrong treatmentLaboratory professionals might believe they are not associated with postanalytic medical errors, but they can be. One deadly example is fatal hemolytic transfusion reaction involving laboratory error.

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Bleeding After a Venipuncture Can Be an Adverse Event

Excessive bleeding after a venipuncture can occur as a preventable or an unpreventable adverse event. Laboratory professionals might or might not have control over this situation because of the factors involved. For example: Bleeding due to failure to apply immediately pressure on the venipuncture site is a preventable adverse event. Bleeding due to later injury to the venipuncture site is an unpreventable event. Circumstances that cause the bleeding determine whether it is a preventable or unpreventable adverse event.

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Sources of Laboratory-Related ErrorsView Page
Human Nature and Error Prevention

The 2000 IOM report states, "It may be part of human nature to err, but it is also part of human nature to create solutions, find better alternatives, and meet the challenges ahead." Everyone should use these positive aspects of human nature to prevent medical errors. Laboratory professionals can do their part by understanding the nature and causes of medical errors and ways to prevent them. They can also benefit by guidance from many clinical, regulatory, and Internet resources that aim to prevent medical errors.

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Medicare Compliance for Clinical Laboratories
Introduction

The government believes that fraud, abuse and waste exist in the healthcare industry today because of cases it has settled and prosecuted.All healthcare providers, including laboratories, make billing errors.The Office of Inspector General (OIG) believes that honest members of the healthcare community can police themselves if they receive guidance.The OIG has published Compliance Program Guidance documents for health care providers, including laboratories.

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What is a voluntary compliance program?

A voluntary compliance program is created by a laboratory based on the OIG's published guidance.It will reduce or eliminate improper billings to Medicare and prevent criminal activities within its company.If a laboratory develops and implements an effective compliance program, it will receive special consideration should a problem arise involving a government investigation.

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Element 7

Element 7: The laboratory has a duty to respond to problems it detects either through its auditing or monitoring system or as reported by employees.It is required to take corrective action and review policies and procedures to ensure the problem does not occur again.It is required to discipline or retrain employees if necessary.The laboratory will report problems to appropriate government agencies and return any money to which it is not entitled.

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Element 1

An effective compliance program includes seven basic elements.Element 1: The laboratory has written and distributed to all affected employees standards, policies and procedures that instruct employees in the proper legal and ethical conduct expected in all areas of business the laboratory conducts.These policies must be adhered to by all employees regardless of status or position in the company.

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Element 2

Element 2: The laboratory has appointed a Compliance Officer (CO) and a Compliance Committee to serve as the focal point for all compliance activities and decision making.The CO and any member of the compliance committee is accessible to all employees in the laboratory.

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Element 3

The laboratory has established a comprehensive training and education program about the laws and regulations that govern the laboratory and the standards, policies and procedures of the compliance program. Mandatory for all employees regardless of status or position in the company. There is additional training for those employees who work in higher compliance risk areas of the laboratory such as billing, marketing, and sales. This interactive software is a component of that training program.

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Element 4

Element 4: The laboratory has established a system of communication so that employees can and are encouraged to report problems and suspect activities they might observe or discover, without fear of retribution, including an anonymous option.The anonymous system instructions are posted throughout the laboratory.

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Element 6

Element 6: The laboratory monitors and regularly audits compliance activities, policies and procedures to ensure they are being followed.If a problem is detected through the audit or monitoring process, it should be reported to the appropriate supervisor, manager or the CO.

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Laws and regulations that govern laboratories

Social Security Act: Medicare and Medicaid laws are in this act. Medicare rules and regulations come under this act. Antikickback laws: Provide criminal penalties for individuals or entities that knowingly and willfully offer, pay, solicit or receive money or favors for referrals of tests or services that will be paid for by the Medicare or Medicaid programs. False Claims Act: Provides criminal penalties for knowingly or willingly filing a false claim to a government program. Self Referral (Stark) laws and regulations: Identify financial relationships that have the potential to result in directed referral to one or both of the individuals or entities involved. Prohibit the referral of patients or tests between related entities unless certain conditions are met. Health Insurance Portability and Accountability Act (HIPAA) Prohibits health care providers and payers from improper or inappropriate use of a patient's confidential health information Requires health care providers to insure that a patient's confidential information is kept secure Provides for standardized electronic formats for all health care transactions

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Risk areas

The government identifies laboratory activities it considers high risk areas for compliance problems.This compliance program focuses on these areas.Training and education for employees lists and explains each of these risk areas.

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Billing and medical necessity

Billing: Highest risk activity a laboratory has. All laboratory activities contribute to the billing process. Many of the risk areas included in this program are components of the billing function. Medical necessity: Medicare is only allowed, by law, to pay for tests that are reasonable and necessary for the diagnosis and treatment of disease. Medical necessity is an underlying principle of the Medicare program. Tests performed for screening or routine exams are not considered medically necessary by the Medicare program.

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Medical coverage policies (LMRPs)

LMRPs (Local Medical Review Policies) are published by Medicare for some laboratory tests. Developed for tests that can be used for screening or diagnosis of disease. CPT codes describe laboratory tests and ICD-9CM codes determine when coverage is allowed. If an LMRP test is ordered by a physician, an ICD-9CM code that is included in the LMRP must be given to the laboratory or the Medicare program will not pay for the test. It is against the law for laboratory to change or add an ICD-9 code submitted by a physician. The Balanced Budget Act of 1997 made it illegal for physicians to order LMRP tests and not supply an ICD-9CM code with the order.

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Advance Beneficiary Notices (ABNs)

Advance Beneficiary Notices (ABNs) allow laboratories to bill Medicare patients directly for specific tests that are not covered by Medicare. A laboratory cannot bill a Medicare Beneficiary for a laboratory test unless it notifies the patient in writing that Medicare is not going to pay for the test. This notice is called an ABN. The beneficiary can choose not have the test performed if they do not want to pay for it. Laboratories cannot make all Medicare beneficiaries sign ABNs. The ABN must contain the specific name of the test. The ABN must give a specific reason the laboratory thinks payment for the test will be denied. The beneficiary should sign the ABN and be given a copy.

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Confidentiality

All employees have a responsibility to maintain the confidentiality of medical information. Medical information should never be discussed outside of the laboratory. It should only be discussed with the ordering doctor or an authorized representative of the doctor. Employees should verify the identity of the individual requesting such information Employees who communicate with patients, physicians or their office staff, insurance company representatives or government employees about any laboratory activity should only give information they know to be true and accurate. Employees should never give false information and should never guess the answer to any question. In case of doubt, refer the person to a supervisor.

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Requisitions and ambiguous orders

Requisitions must be designed to ensure that ordering physicians can choose tests that are medically necessary for their patients. Requisitions should contain reminders about Medicare rules of medical necessity and list the contents of panels and profiles. Requisitions must provide a place for the physician to include diagnosis (ICD9-CM) codes. Physicians should be encouraged to use only the requisitions supplied by the laboratory to order tests. Ambiguous or unclear test orders When the orders for a test are not absolutely clear, the laboratory must contact the ordering physician to clarify the orders before performing and billing for the test. The laboratory cannot guess at the order. The laboratory cannot perform and bill for tests that are not specifically ordered. The laboratory cannot change a physician order without contacting the physician. In any case where specimen integrity or patient care will be compromised by a delay in testing follow the policies the laboratory has established for such cases.

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Tests performed and ordered correctly

The laboratory has a system in place to detect tests that are not performed due to a laboratory error and stop or credit the billing for these tests. The laboratory cannot bill for tests that are not performed. Employee aware of a test being canceled or not being performed for some reason must follow the policies and procedures associated with correcting the billing. The laboratory only performs tests that are ordered by individuals authorized to order tests. If an employee knows that a test has been ordered by someone other then an authorized individual, the employee should report it to their supervisor or the CO.

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Panels and profiles

Panels and Profiles: It is not against the law for a laboratory to allow the use of panels, profiles and custom panels. The laboratory must ensure that the ordering doctor knows what tests are included in a panel or profile and what CPT codes will be billed to the Medicare program. The laboratory notifies doctors about panels and profiles through a written notice and the requisition. Employees should not permit the order of any panel or profile not authorized by the laboratory.

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Physician notices and acknowledgements

Notices to physicians must be sent by the laboratory to its customers once each year. Notices remind physicians about Medicare rules and regulations. Notices include summaries of laboratory test ordering policies, requisition use, CPT coding and ICD coding. Physician acknowledgements must be signed by any physician who wants to create a custom panel, profile or reflex test. This is the only way a special panel or profile may be performed by the laboratory. The physician must order tests individually when there is no physician acknowledgement signed. The laboratory must renew physician acknowledgements at least annually.

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Inducements

It is against the law to offer or ask for money or favors to get a physician to order tests from a laboratory. This is known as an "inducement" or a "kickback." Laboratories should only give supplies to a physician for the drawing, processing, storing or transporting of specimens to the laboratory. The laboratory cannot provide supplies physicians use for their own purposes. The laboratory must monitor the amount of supplies provided to ensure that it matches the number of tests sent to the laboratory. The lab cannot give free tests except in the event of laboratory error. The lab cannot give free education to clients unless it is about the laboratory's services or policies. The lab cannot give excessive or expensive gifts or entertainment to physicians The lab can give discounts but the price must be above cost and at "fair market value."

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Phlebotomists and equipment in client offices

Laboratories may place phlebotomists or other employees in a physician office if all of the following are done: Employee only performs laboratory related tasks. There is a written understanding given to the physician about what the employee can and cannot do. Periodic audits are done to ensure the employee is following these policies. Laboratories may place printers, computers, fax machines or other equipment or products in client offices as long as they ensure that: The physician understands the equipment belongs to the laboratory. It is used for laboratory purposes like receiving reports or ordering tests. Periodic audits are done to ensure that the client is using the equipment only for laboratory related activities.

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Couriers and referral tests

The laboratory's couriers may not transport items except those related to the testing services offered by the laboratory. Couriers must follow all OSHA standards for the handling and transport of specimens. The laboratory is responsible for all tests it refers to other laboratories. Laboratory should not change CPT codes supplied by a reference laboratory without contacting the reference laboratory. The laboratory is responsible for all tests it bills to Medicare/Medicaid even if the test was performed by a reference laboratory.

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Marketing and record retention

The laboratory must ensure that its sales and marketing materials are clear and not deceptive: They should fully inform the physician about the appropriate use of laboratory tests and services. They should not be designed to induce unnecessary testing. The laboratory must have in place a record retention policy that ensures records are accessible in case of an audit or investigation. Records should be retrievable. Related documents should be linked.

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Utilization and other regulations

Laboratories must not induce physicians to order unnecessary tests through their marketing or education activities: They must monitor the use of laboratory services by their clients. They must correct any situation where something they did caused an unnecessary increase in test utilization. Cost Reports Hospitals laboratories must ensure that information used in hospital Medicare cost reports is accurate and includes only those costs which are appropriate. Laboratories must follow all CLIA and OSHA regulations: failure to do so may result in a False Claims Act violation.

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Billing

Billing is the highest risk area for the laboratory because it generates the claims that are sent to Medicare and other government payers.Payment and payment errors are the focus of the OIG compliance guidance for the laboratory because of the revenue involved.Fraud and abuse is often perpetrated as a billing scheme.Nearly all laboratory functions can affect the billing of laboratory tests.

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Medical necessity

Medical necessity means that Medicare is not allowed by law to pay for any tests that are not necessary for diagnosis or treatment of disease.A laboratory may not submit a claim to Medicare or other government payers for any test it knows is not medically necessary except in certain cases: When the patient has signed an advance notice. When a patient has requested the lab to submit such a claim for a determination by Medicare. Medicare does not pay for screening tests or tests that are ordered in the absence of signs or symptoms.Billing department employees are responsible to follow all policies and procedures related to the submission of claims to reduce erroneous billings.

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HCPCS and CPT coding

The HCFA (Health Care Financing Administration) Common Procedural Coding System (HCPCS) and the CPT (Current Procedural Terminology) codes are used to describe specific tests or services. The amount of payment for a test is dependent on the HCPCS or CPT code. HCPCS or CPT codes should be assigned under the supervision of the laboratory technical staff. Billing department employees should never change a HCPCS or CPT code without the approval of a manager or compliance officer. If a billing department clerk notices that a particular HCPCS or CPT code is being rejected by a payer they should report it to their manager. It is against the law to use the wrong HCPCS or CPT code for the purpose of causing or increasing payment for a test.

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ICD-9CM coding

ICD-9CM (International Classification of Disease, 9th Edition, Clinical Modification) codes are used for the classification of disease and conditions and for describing signs, symptoms and medical circumstances.These codes are used to indicate the medical necessity of a particular test.ICD-9 codes can only be supplied by the ordering physician or a representative of that physician. "Code steering" means to steer or direct a physician to supply an ICD-9 code that is payable. ICD-9 codes cannot be used from a previous laboratory order. If a physician supplies a narrative description instead of an ICD-9 code the laboratory must accurately translate that code using only certified coders.It is against the law to use the wrong ICD-9 code for the purpose of causing or increasing payment for a test.

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Local medical review policies (LMRPs)

LMRPs (Local Medical Review Policies) are published by Medicare for some laboratory tests. They are usually developed for tests that can be used for screening or diagnosis of disease. LMRPs use CPT codes to identify the tests and ICD-9 codes to determine when coverage is allowed. If an LMRP test is ordered by a physician, an ICD-9 code that is included in the LMRP must be given to the laboratory or the Medicare program will not pay for the test. It is against the law for laboratory to change or add an ICD-9 code submitted by a physician. A laboratory should not submit a claim for an LMRP test that is not accompanied by an acceptable ICD-9 code. The Balanced Budget Act of 1997 made it illegal for physicians to order LMRP tests and not supply an ICD-9CM code with the order.

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Communication with physicians and patients

It is important that billing department employees are clear and accurate when communicating with physician office personnel and patients. Never guess at the answer to a question; ask if you are unsure. Do not speculate or express personal opinions. When requesting diagnosis information from the physician office staff be careful to not lead them to give a billable code: The code must come from the patient's medical record. There is an incentive program for patients to find and report fraud and abuse by health care providers, including laboratories, so: Billing department employees must accurately state laboratory policies and procedures, or forward the call to a supervisor to avoid misstatements and misunderstandings.

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Advance Beneficiary Notices (ABNs)

A Laboratory may not bill a Medicare Beneficiary for a test unless it notifies the patient in writing before the testing is done that Medicare is not going to pay for the test. This notice is called an ABN. Laboratories cannot make all Medicare beneficiaries sign ABNs. The ABN must contain the specific name of the test and give a specific reason the laboratory thinks payment for the test will be denied. The beneficiary should sign the ABN and a copy should be sent to the laboratory and one given to the beneficiary. The billing department must have evidence that the ABN has been signed before it bills a patient.A laboratory may bill Medicare even though it knows it will not be paid when it has evidence an ABN has been signed. A modifier (GA) must be added to the CPT code for a test where an ABN has been signed.

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Excused charges and other inducements

The laboratory should not offer or provide free testing to any individual in a position to make or control referral for laboratory services: The laboratory may write off charges only when laboratory errors in billing or testing occur. Sales and marketing personnel cannot offer free testing in any form unless approved by the compliance officer. Free testing for indigent patients must be approved by the compliance officer. Sales and marketing personnel cannot offer or give anything of value to a customer or potential customer beyond the usual promotional items. If a client solicits a questionable or illegal item or special consideration, it should be reported to the sales manager or compliance officer.

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Test pricing and antitrust

It is not against the law for a laboratory to have different fee schedules for different billing situations.Most laboratories have one fee schedule for customers that must be billed individually (patients, insurance, Medicare) and one for customers billed monthly on an invoice type of statement (client or doctor billing).The difference in price between the two schedules should be a reflection of the financial benefits of direct client billing.Test prices should be determined by means of a financial analysis that include such factors as cost, market value and reasonable profit.Contractually arranged pricing that results from negotiations with insurance and managed care companies should at least cover costs of testing.Laboratories may not work together to fix or set prices in the market place.

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Equipment and space

Laboratories may only lease space from physicians who refer Medicare patients to them under certain circumstances: There must be a written lease for at least one year. Lease price must be at "fair market value." All leases must be reviewed by legal counsel to ensure compliance with antikickback and Stark laws.When leasing or renting equipment to a physician or from a physician the same basic rules apply as for space.If the laboratory is located in a hospital, the relationship between the hospital and a physician who refers to the lab may have antikickback or Stark implications.

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Client contracts

A laboratory that receives referrals from a nursing home or Skilled Nursing Facility (SNF) should have a written agreement with that facility: The Agreement should define billing and documentation responsibilities. The facility should be responsible for determining the payment status of its patients and is liable for submitting incorrect payment information to the laboratory. Fees should be consistent with other similar customers. A laboratory that provides services to a Home Health Agency treating Medicare/Medicaid beneficiaries should have a written agreement with that agency: The Agreement should define billing and documentation responsibilities. The Agreement should place the responsibility on the Home Health Agency to establish that all patients receiving laboratory services are "homebound" as defined by Medicare. Fees should be consistent with other similar customers.

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ICD-9 codes and ABNs

Never use sign and symptom information received from a patient for laboratory billing purposes.Never use ICD-9 codes from previous visits for a new visit even if the request is for the same test and the patient assures you that it is for the same reason. (Standing orders are an exception.)ICD-9 codes should be requested when setting up standing orders and will then apply to all subsequent visits. It is not necessary in this case to get a new ICD-9 code for each visit.If the patient refuses to sign an ABN but demands to have the test done: Have the fact that they were given notice (ABN) witnessed by a second person. (By phone if you are located in single-person drawing site). Ensure that documentation is complete.

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Confidentiality and inducements

Do not leave test orders or test results in areas where they can be viewed by patients.Do not discuss test results or any patient information in areas where patients can overhear the conversation. Be careful not to discuss confidential information on the telephone where patients can overhear the conversation.Do not provide supplies to physician offices other than those usually provided by the laboratory. Document any supplies given to an office.Do not supply items that the office can use for testing (e.g. urine dipsticks). Do not allow offices to dispose of biohazard waste or sharps in the waste containers paid for by the laboratory.

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Communicating with patients in person

When communicating directly with patients concerning their laboratory tests or orders be careful not to discuss the following: Why a test has been ordered by a physician. What the test might indicate or what the test results mean. Any opinions about their doctor. Any information about internal laboratory issues. Refer the patient to their doctor for information about the tests and the results.If the laboratory provides any printed information about tests that are designed for patients, give the patient that information.

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Test orders

Anytime an order is not clear, the physician office must be contacted.Do not use information supplied by a patient to clarify an order. Patients cannot add tests on their own. If a patient insists they want tests not specifically ordered by the doctor, the doctor should be contacted.When transferring a doctor's order from a non-standard form like a prescription pad to a laboratory requisition, it is important to ensure the accuracy of the order.Attach the original order document to the requisition sent to the laboratory.Follow all laboratory policies about panels and profiles, ambiguous orders and reflex tests.

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Case Study 9

The setting is automated chemistry department, night shift, busy core laboratory for a hospital based outreach laboratory. A medical technologist who operates the automated chemistry analyzer on third shift encounters short samples a couple of times a night. When this happens, he runs as many of the ordered tests as he can and fills in the blank results with a comment indicating that a short sample occurred. As far as he knows there isn't a policy that addresses this problem directly.The test reports out with the results and the comments. The technologist does not have to change the physician order in any way and is providing the maximum results that can be reported for the specimen in a timely fashion. This is done as a matter of patient care and quality service. There has not ever been a complaint about this practice as far as he knows. Are there any additional steps this technologist should be taking?Correct Answer: The technologist should follow the procedures that the laboratory has in place for testing and billing samples for which there is no order or for ambiguous orders. If the policies do not seem to address his particular situation, he thinks there should be a separate policy to cover this situation or has a question about it, he should talk to his supervisor or to the laboratory compliance officerDiscussion: This choice addresses the problem in the most complete manner, in that the employee fulfills his responsibility to take action when he thinks there is a problem.

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Case Study 10

The setting is nursing home where a phlebotomist from the laboratory goes to draw blood samples each day. The phlebotomist picks up the requisitions for blood test orders at the nursing station and then goes to the various rooms to draw blood from the patients. She notices that every requisition has an Advanced Beneficiary Notice (ABN) attached to it that is signed by the patient, even when the tests that were ordered don't need them. She asks the nurse at the station but she informs the phlebotomist that she doesn't know anything about it because it is done on the night shift.She lets the phlebotomist know that she will inform the nursing supervisor about it when she arrives at 9:00 AM. The phlebotomist completes her blood draws and returns to the laboratory. What should the phlebotomist do, if anything, in addition to her letting the nurse know about the problem?Correct Answer: The phlebotomist should report the incident to her supervisor upon returning to the laboratory.Discussion: Since the laboratory is submitting the claims for any Medicare patients that the phlebotomist might draw, the problem is the labs problem. However, it is not going to change the fact that the ABNs were already signed by the patients if the phlebotomist refuses to draw them or if the nursing personnel are required to remove them. By contacting the supervisor, an appropriate representative from the laboratory can follow up with the nursing supervisor to ensure they understand the laws and regulations that govern ABNs.

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Case Study 7

The setting is the cafeteria in a hospital or the lounge in an independent laboratory. Two employees from different departments are old friends are having lunch together. A billing clerk and a medical technologist are friends and are having lunch together. The billing clerk mentions that she saw a bill go through the system for one of her coworkers for a biopsy. She asks the medical technologist if she has the necessary security level access to see pathology test results because she is concerned about the welfare of the coworker. The medical technologist does have the necessary security clearance to see the results. She should:Correct Answer: Refuse to look up the results for the clerk and remind the clerk that it is a violation of compliance policies to do so, or to ask another to do so. Remind her of the requirement for each employee to report any violations of policy. Discussion: The Medical technologist has a responsibility to report violations of compliance policies and the friend has put her in a difficult position. For that reason, it is not enough to just refuse the clerk's request. If the medical technologist does not take the responsibility to inform the employee of the policy then there is a possibility that the employee would ask some other employee to do it for her.

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Case Study 8

The setting is a billing office in a laboratory where two or more clerks work together in very close proximity with each other, so that each can easily see what the other is doing. A billing clerk notices that one of his fellow employees is changing or adding codes to requisitions he is processing. This employee is a friend of his and he knows that he really needs the job at the laboratory because he is a single parent raising two kids. He also knows that what the employee is doing is against the company's compliance policies.He asks the employee about it and is given the explanation that because the computer requires something to be entered in the ICD-9 code field and he only does this with non-Medicare patients, it doesn't matter. The employee explains that it saves him a lot of time he uses to call to get codes for the Medicare patients. What should this clerk do about this?Correct Answer: She should talk to the supervisor about the problem even if she talks to the employee about it and the employee says she will talk to the supervisor and stop doing it.Discussion: Every employee who becomes aware of a violation of the law or a compliance policy has a responsibility to take action, which includes reporting the problem to a supervisor or the compliance officer. It doesn't matter that these patients are not Medicare patients, the important thing is that the employee is violating a compliance policy. If this employee does not report the problem he is himself violating a compliance policy. If it is subsequently discovered that he knew and didn't report it, he could be terminated. If there is a need for refunds to be done or other action, it will not occur and could create a big problem for the lab in a subsequent audit or other action.

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Case Study 5

The Client Services department which is a crowded room divided into cubicles which contain desks separated only by thin moveable dividers. Lots of activity, phones ringing, multiple conversations going on at once etc. A client service representative receives a call from a large client office that she speaks with every day for a variety of reasons. Today the client is requesting the laboratory to write off the charges for a test that the office person ordered incorrectly by mistake even though the laboratory has already done the test and reported the results back to the office. Since this service representative works with this office frequently she believes that this is a rare request. Actions that the client service representative may take are:Correct Answer: Refuse to write off the charges for the test and inform the client that it could be considered an inducement if the laboratory does that, which would make both the laboratory and the office liable should it ever come to light. Offer whatever billing options are available according to lab policies. or Refuse to write off the charges and explain to the client that approval must be obtained from the department manager or the laboratory compliance officer before any action can be taken because writing the test off could be considered an inducement.Discussion: The primary reason is that the test is not written off simply because the client asks for it to be done. Tests should never be written off by the laboratory automatically. There should always be an approval process involved or a policy that strictly forbids any write off except in the case an error on the part of the laboratory where documentation exists to support it.

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Case Study 6

The setting is a large outreach or independent laboratory that processes a high volume of tests with tight resources and a lot of customer service turn around time deadlines to meet that are dependent on the specimen processing department to complete their work on time, every day and night. The manager of the specimen processing department receives a memorandum from the compliance officer (CO) that several of his employees have not attended their compliance training sessions.Since this is the initial training for the laboratory, the CO reminds the manager of the requirement that all employees attend. The employees listed in the memo are key employees, who are top performers in the department. The manager knows that they have been instructed to attend, and that attendance was mandatory but left it up to them to choose the session they would attend. When questioned about their non attendance, all of the individuals said they were too busy to attend and it was a matter of going to the classes or getting the work done. What action should the manager take in this situation?Correct Answer: In consideration of the fact that these are the initial training sessions, sit down with each of the affected employees and very specifically tell them that the classes are mandatory and that they must attend under penalty of disciplinary action.Discussion: The manager knows that the compliance policy is mandatory attendance for these training sessions and informed these employees of that fact. When taking the disciplinary action, the manager may take into account the fact that these employees have not received the training and therefore may not understand the implications of not attending, but the some appropriate discipline should be taken. The manager should also give some consideration to the mentality that caused these employees to skip the training in the first place even after being instructed to attend. This same kind of decision process, the work must get done, is a common cause of compliance problems occurring.

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Case Study 2

A courier is making a routine stop in a client's office and is approached by the office manager with whom he is very friendly because he has been going to this office for years. The office manager asks the courier if Dr. John Smith is a regular stop on his route and the courier answers yes. She then asks the courier if he would do her a little favor since he stops at Dr Smiths office regularly anyway and drop off an x-ray for her so she won't have to call a courier service. The courier knows that this is a big account for the laboratory and customer service is a high priority for the laboratory. This courier should:Correct Answer: Refuse to do it for the customer and explain to the customer that the laboratory has a policy that says he must only provide courier services related to laboratory.Discussion: Even though this is a single incident, by doing this favor the courier is giving the office manager license to ask these kinds of favors in the future. Since the provision of this free courier service is a form of inducement or kickback to the client, both the client and the laboratory would be involved if the such a practice were to go on regularly and be discovered by the government or by the laboratory. Hiding the incident and asking the office manager to conspire with him to do this will only make it worse for the employee and would lead to serious discipline action up to termination. The courier's best course of action, for the protection of his friend the office manager and himself, the physician practice and the laboratory is to not do this and explain the reason to the office manager so she is aware of the consequences of asking this favor.

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Case Study 3

It is 11:00 PM and the specimen processing department is finishing up the night's accessioning and test requesting. A specimen processor is working on a requisition that has an order for a Hepatic Profile but there are two tubes of blood with the order, one of which is a lavender top tube. This is the fourth requisition from this same doctor's office and all of them have had a lavender top tube and serum tube with an order for a chemistry test and a CBC. No CBC is marked on the requisition or written on the tube. The specimen processor figures the office just forgot to mark the test and knows that the results will be delayed and the sample might not be any good if he doesn't order the CBC now. He is also under pressure from the technical departments to finish processing on time so they can get their work done on time for result printing in the morning. What should the processor do?Correct Answer: Look up the laboratory's policy for handling such a situation and follow the policy.Discussion: The laboratory is not permitted to change a doctor's order in any way. By ordering the CBC the processor is ordering a test that the doctor did not specifically order and therefore makes the laboratory subject to a violation of the False Claims Act. By reviewing and following the laboratory policy the processor assures that the laboratory, the physician and the patient's best interests are met.

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Case Study 4

Busy hospital laboratory in a 350 bed urban hospital that provides laboratory testing for the hospital and for the hospital's outreach testing laboratory. A medical technologist in the microbiology department receives a call from a friend who works in a laboratory in a physician office. The physician is not a regular client of the laboratory currently but uses another laboratory for most of their work. The microbiologist knows that the sales department would like to get this account. The friend explains to her that she is doing a quality control check on her in-office microbiology testing and her regular laboratory will do it but is going to charge her for it. She asks the microbiologist if she will do it for free since it is quality control, not Medicare and is not going to be billed to anyone.She tells the microbiologist that she would like to use the hospital lab for everything but her doctor insists on using the competitor. She indicates that the favor might help get the doctor to try the hospital laboratory for other tests. The microbiologist should:Correct Answer: Explain to her friend that if the hospital does the tests for no charge on the promise of other referrals, both the physician office and the hospital could be liable for violations of the antikickback statute.Discussion: The antikickback statute is implicated in this scenario because the free testing is solicited on the condition that other referrals may occur as a result of providing the favor. In fact, the solicitation itself is a violation of the law. The fact that Medicare patients are not specifically mentioned in the scenario is not sufficient to remove the risk. The technologist should also report the incident to the Compliance Officer and seek advise about what documentation, if any, should be kept concerning the incident.

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Case Study 1

A billing clerk is entering billing demographics on requisitions as a part of the normal days work. The department is under pressure to reduce accounts receivable, which means that the more clean claims that are filed, the better. This particular requisition is for a Medicare patient and has an LMRP test but does not have a diagnosis on it. She remembers that just a few requisitions before this one she had a requisition from the same doctor that had this same test on it that did have a diagnosis that allowed the test to be billed. She thumbs back in the pile and finds the previous requisition, notes the code that was used and adds it to the current requisition. This will help her meet the department goal of getting claims paid and reducing accounts receivable. It is all right for her to do this because:Correct Answer: She should not do this because it is against the law to change diagnosis information on a requisition.Discussion: A laboratory employee should never change, add or use previously received diagnosis information for the purpose of making a test billable for the Medicare program or for any other insurance or payer. This is a form of fraud and for each claim submitted as a result of this activity the laboratory is liable for a false claim and would have to pay the government back three up to times the reimbursement for the test and up to $10,000 for each claim submitted. Further, if the employee is caught doing this, even if the employee is ignorant of the law and any laboratory policy prohibiting it, she must be disciplined and so should the supervisor. Any employee who notices another employee doing this should correct the employee and report the incident to the department supervisor immediately.

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Company Responsibility

The company also carries certain responsibilities: The laboratory has a responsibility to communicate to all its employees its compliance policies, procedures, standards of conduct and the consequences of non-compliance. All employees must be able to understand the policies and standards. The laboratory has a responsibility to act on reported problems and suspect activities. The laboratory must resolve problems and ensure they don't recur. The laboratory must take appropriate and fair disciplinary actions against all involved. The laboratory must report to the appropriate government agency when necessary. The laboratory must promptly return money received from the government to which it is not entitled. The laboratory must audit its policies and procedures to ensure they are being followed and that they work.

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Employee's Responsibility

An employee has certain responsibilities: Voluntary Compliance Programs should detect and prevent billing errors and fraud in the clinical laboratory. Compliance programs must be effective to be beneficial in case of an investigation. Compliance is every employee's responsibility regardless of status or position in the company. All employees are subject to disciplinary action if they violate compliance policies or laws. Employees must understand their responsibility to report any problems or suspect activity they encounter in the laboratory. Employees will not be disciplined for reporting problems. Employees should use the Hotline or other established anonymous reporting system if they are afraid of retribution.

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Medicare Compliance for Clinical Laboratories (updated 2009)
Laboratory Administration's Responsibilities

Laboratory administration has these responsibilities: Communicate compliance policies, procedures, standards of conduct to all employees and the consequences of non-compliance. Act on reported problems and suspect activities. Resolve problems and ensure they don't recur. Take appropriate and fair disciplinary actions against all involved. Report to the appropriate government agency when necessary. Promptly return money received from the government to which the laboratory is not entitled. Audit the laboratory's policies and procedures to ensure they are being followed and that they work.

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Employee's Responsibility

Compliance is every employee's responsibility regardless of status or position in the company. All employees are subject to disciplinary action if they violate compliance policies or laws. Employees must understand their responsibility to report any problems or suspect activity they encounter in the laboratory. Employees will not be disciplined for reporting problems. Employees should use the Hotline or other established anonymous reporting system if they are afraid of retribution.

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What is a voluntary compliance program?

All healthcare providers, including laboratories, could potentially submit erroneous claims for Medicare reimbursement. These billing errors can trigger an investigation. The creation of a voluntary compliance program, using the guidelines provided by the Office of Inspector General (OIG), can assist healthcare institutions and laboratories to audit themselves, thereby preventing submission of erroneous claims and a possible fraud and abuse investigation.

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Element 7

The laboratory has a duty to respond to problems it detects either through its auditing or monitoring system or as reported by employees. It is required to take corrective action and review policies and procedures to ensure the problem does not occur again. It is required to discipline or retrain employees if necessary. The laboratory will report problems to appropriate government agencies and return any money to which it is not entitled.

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Element 1

The laboratory has written and distributed to all affected employees, standards, policies, and procedures that instruct employees in the proper legal and ethical conduct expected in all areas of business the laboratory conducts. These policies must be adhered to by all employees regardless of status or position in the company.

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Element 2

The laboratory has appointed a Compliance Officer (CO) and a Compliance Committee to serve as the focal point for all compliance activities and decision making. The CO and any member of the compliance committee is accessible to all employees in the laboratory.

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Element 3

The laboratory has established a comprehensive training and education program about the laws and regulations that govern the laboratory and the standards, policies and procedures of the compliance program. Mandatory for all employees regardless of status or position in the company. There is additional training for those employees who work in higher compliance risk areas of the laboratory such as billing, marketing, and sales. This interactive software is a component of that training program.

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Element 4

The laboratory has established a system of communication so that employees can, and are encouraged to, report problems and suspect activities they might observe or discover, without fear of retribution, including an anonymous option. The anonymous system instructions are posted throughout the laboratory.

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Element 6

The laboratory monitors and regularly audits compliance activities, policies, and procedures to ensure they are being followed. If a problem is detected through the audit or monitoring process, it should be reported to the appropriate supervisor, manager or the CO.

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Test Requisitions

Requisitions must be designed to ensure that ordering physicians can choose tests that are medically necessary for their patients. Requisitions should contain reminders about Medicare rules of medical necessity and list the contents of panels and profiles. Requisitions must provide a place for the physician to include diagnosis (ICD9-CM) codes. Physicians should be encouraged to use only the requisitions supplied by the laboratory to order tests.

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Issues Related to Test Ordering, Performance, and Resulting

The laboratory can only perform tests that are ordered by individuals authorized to order tests. If an employee knows that a test has been ordered by someone other then an authorized individual, the employee should report it to their supervisor or the compliance officer (CO). The laboratory must have a system in place to detect tests that are not performed due to a laboratory error and stop or credit the billing for these tests. The laboratory cannot bill for tests that are not performed. An employee who is aware that a test has been canceled or has not been performed for some reason must follow the policies and procedures associated with correcting the billing. Release of test results by phone, fax and other non-routine methods Employees should release test results only to the clinical person who is authorized to receive the test results (i.e., the ordering physician). Never release test results to another physician or entity unless authorized by the ordering physician in writing. Never release the results of a test to a patient unless authorized in writing by the ordering physician.

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Ambiguous or Unclear Test Orders

When the orders for a test are not absolutely clear, the laboratory must contact the ordering physician to clarify the orders before performing and billing for the test. Do not attempt to guess what the order is. The laboratory cannot perform and bill for tests that are not specifically ordered. The laboratory cannot change a physician order without contacting the physician. In any case where specimen integrity or patient care will be compromised by a delay in testing, follow the policies the laboratory has established for such cases.

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Panels and profiles

Panels and Profiles: It is not against the law for a laboratory to allow the use of panels, profiles and custom panels. However, all applicable CPT codes must be included. The laboratory must ensure that the ordering doctor knows what tests are included in a panel or profile and what CPT codes will be billed to the Medicare program. The laboratory notifies doctors about panels and profiles through a written notice and the requisition. Employees should not permit the order of any panel or profile not authorized by the laboratory.

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Physician notices and acknowledgements

Notices must be sent annually by the laboratory to its customers (physicians). Notices remind physicians about Medicare rules and regulations. Notices include summaries of laboratory test ordering policies, requisition use, CPT coding and ICD coding. Physician acknowledgements must be signed by any physician who wants to create a custom panel, profile or reflex test. This is the only way a special panel or profile may be performed by the laboratory. The physician must order tests individually when there is no physician acknowledgement signed. The laboratory must renew physician acknowledgements at least annually.

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Excused charges and other inducements

The laboratory should not offer or provide free testing to any individual in a position to make or control referral for laboratory services: The laboratory may write off charges only when laboratory errors in billing or testing occur. Sales and marketing personnel cannot offer free testing in any form unless approved by the compliance officer. Free testing for indigent patients must be approved by the compliance officer. Sales and marketing personnel cannot offer or give anything of value to a customer or potential customer beyond the usual promotional items. If a client solicits special consideration, it should be reported to the sales manager or compliance officer.

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Test pricing and antitrust

It is not against the law for a laboratory to have different fee schedules for different billing situations.Most laboratories have one fee schedule for customers that must be billed individually (patients, insurance, Medicare) and one for customers billed monthly on an invoice type of statement (client or doctor billing).The difference in price between the two schedules should be a reflection of the financial benefits of direct client billing.Test prices should be determined by means of a financial analysis that include such factors as cost, market value and reasonable profit.Contractually arranged pricing that results from negotiations with insurance and managed care companies should at least cover costs of testing.Laboratories may not work together to fix or set prices in the market place.

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Client contracts

A laboratory that receives referrals from a nursing home or Skilled Nursing Facility (SNF) should have a written agreement with that facility: The Agreement should define billing and documentation responsibilities. The facility should be responsible for determining the payment status of its patients and is liable for submitting incorrect payment information to the laboratory. Fees should be consistent with other similar customers. A laboratory that provides services to a Home Health Agency treating Medicare/Medicaid beneficiaries should have a written agreement with that agency: The Agreement should define billing and documentation responsibilities. The Agreement should place the responsibility on the Home Health Agency to establish that all patients receiving laboratory services are "homebound" as defined by Medicare. Fees should be consistent with other similar customers.

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Case Study 9

The setting is a nursing home where a phlebotomist from the laboratory goes to draw blood samples each day. The phlebotomist picks up the requisitions for blood tests at the nursing station and then goes to the various rooms to draw blood from the patients. She notices that every requisition has an Advanced Beneficiary Notice (ABN) attached to it that is signed by the patient, even when the tests that were ordered don't need them. She asks the nurse at the station but she informs the phlebotomist that she doesn't know anything about it because it is done on the night shift.She lets the phlebotomist know that she will inform the nursing supervisor about it when she arrives at 9:00 AM. The phlebotomist completes her blood draws and returns to the laboratory. What should the phlebotomist do, if anything, in addition to her letting the nurse know about the problem?Correct Answer: The phlebotomist should report the incident to her supervisor upon returning to the laboratory.Discussion: Since the laboratory is submitting the claims for any Medicare patients that the phlebotomist might draw, the problem is the lab's problem. However, it is not going to change the fact that the ABNs were already signed by the patients if the phlebotomist refuses to draw them or if the nursing personnel are required to remove them. By contacting the supervisor, an appropriate representative from the laboratory can follow up with the nursing supervisor to ensure they understand the laws and regulations that govern ABNs.

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Case Study 7

The setting is the cafeteria in a hospital or the lounge in an independent laboratory. Two employees, a billing clerk and a medical technologist, from different departments are having lunch together. The billing clerk mentions that she saw a bill go through the system for one of her coworkers for a biopsy. She asks the medical technologist if she has the necessary security level access to see pathology test results because she is concerned about the welfare of the coworker. The medical technologist does have the necessary security clearance to see the results. She should:Correct Answer: Refuse to look up the results for the clerk and remind the clerk that it is a violation of compliance policies to do so, or to ask another to do so. Discussion: The medical technologist has a responsibility to report violations of compliance policies and the friend has put her in a difficult position. For that reason, it is not enough to just refuse the clerk's request. If the medical technologist does not take the responsibility to inform the employee of the policy then there is a possibility that the employee would ask some other employee to do it for her.

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Case Study 8

The setting is a billing office in a laboratory where clerks work in close proximity to each other, so that each can easily see what the other is doing. A billing clerk notices that one of his fellow employees is changing or adding codes to requisitions he is processing. This employee is a friend of his and he knows that he really needs the job at the laboratory because he is a single parent raising two kids. He also knows that what the employee is doing is against the company's compliance policies.He asks the employee about it and is given the explanation that because the computer requires something to be entered in the ICD-9 code field and he only does this with non-Medicare patients, it doesn't matter. The employee explains that it gives him more time to call for codes for the Medicare patients. What should this clerk do about this situation?Correct Answer: He should talk to the supervisor about the problem even if he talks to his fellow employee about it and the employee says he will stop doing it.Discussion: Every employee who becomes aware of a violation of the law or a compliance policy has a responsibility to take action, which includes reporting the problem to a supervisor or the compliance officer. It doesn't matter that these patients are not Medicare patients, the important thing is that the employee is violating a compliance policy. If this employee does not report the problem he is himself violating a compliance policy. If it is subsequently discovered that he knew and didn't report it, he could be terminated. The employee's addition of the codes could create a big problem for the lab iif an audit or inspection occurred.

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Case Study 5

The Client Services department is a crowded room divided into cubicles which contain desks separated only by thin moveable dividers. Lots of activity is present including phones ringing, multiple conversations going on at once, etc. A client service representative receives a call from a large client office that she speaks with every day for a variety of reasons. Today the client is requesting the laboratory to write off the charges for a test that the office person ordered by mistake, even though the laboratory has already done the test and reported the results back to the office. Since this service representative works with this office frequently, she believes that this is a rare request. Actions that the client service representative may take are:Correct Answers: Refuse to write off the charges for the test and inform the client that it could be considered an inducement if the laboratory does that, which would make both the laboratory and the office liable should it ever come to light. Offer whatever billing options are available according to lab policies. or Refuse to write off the charges and explain to the client that approval must be obtained from the department manager or the laboratory compliance officer before any action can be taken because writing the test off could be considered an inducement.Discussion: Tests should never be written off by the laboratory automatically. There should always be an approval process involved or a policy that strictly forbids any write off except in the case of an error on the part of the laboratory where documentation exists to support it.

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Case Study 6

The setting is a large outreach or independent laboratory that processes a high volume of tests with tight resources and a lot of customer service turn around time deadlines to meet that are dependent on the specimen processing department to complete their work on time. The manager of the specimen processing department receives a memorandum from the compliance officer (CO) that several of his employees have not attended their compliance training sessions.Since this is the initial training for the laboratory, the CO reminds the manager of the requirement that all employees attend. The employees listed in the memo are key employees who are top performers in the department. The manager knows that they have been instructed to attend, and that attendance is mandatory, but the manager had left it up to the employees to choose the session they would attend. When questioned about their non-attendance, all of the individuals said they were too busy to attend and it was a matter of going to the classes or getting the work done. What action should the manager take in this situation?Correct Answer: In consideration of the fact that these are the initial training sessions, sit down with each of the affected employees and very specifically tell them that the classes are mandatory and that they must attend under penalty of disciplinary action.Discussion: The manager knows that the compliance policy is mandatory attendance for these training sessions and informed these employees of that fact. The manager may take into account the fact that these employees have not received the training and therefore may not understand the implications of not attending, but some appropriate discipline should be taken. The manager should also give some consideration to the mentality that caused these employees to skip the training in the first place even after being instructed to attend. This same reasoning that "the work must get done", is a common cause of compliance problems.

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Case Study 2

A courier is making a routine stop in a client's office and is approached by the office manager with whom he is very friendly because he has been going to this office for years. The office manager asks the courier if Dr. John Smith is a regular stop on his route and the courier answers yes. She then asks the courier if he would do her a little favor since he stops at Dr Smiths office regularly anyway and drop off an x-ray for her so she won't have to call a courier service. The courier knows that this is a big account for the laboratory and customer service is a high priority for the laboratory. This courier should:Correct Answer: Refuse to do it for the customer and explain to the customer that the laboratory has a policy that says he must only provide courier services related to laboratory.Discussion: Even though this is a single incident, by doing this favor the courier is giving the office manager license to ask these kinds of favors in the future. Since the provision of this free courier service is a form of inducement or kickback to the client, both the client and the laboratory would be involved if the such a practice were to go on regularly and be discovered by the government or by the laboratory. Hiding the incident and asking the office manager to conspire with him to do this will only make it worse for the employee and would lead to serious disciplinary action up to termination.

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Case Study 3

It is 11:00 PM and the specimen processing department is finishing up the night's accessioning and test order entry. A specimen processor is working on a requisition that has an order for a Hepatic Profile but there are two tubes of blood with the order, one of which is a lavender top tube. This is the fourth requisition from this same doctor's office and all of them have had a lavender top tube and serum tube with an order for a chemistry test and a CBC. No CBC is marked on the requisition or written on the tube. The specimen processor figures the office just forgot to mark the test and knows that the results will be delayed and the sample might not be any good if he doesn't order the CBC now. He is also under pressure from the technical departments to finish processing on time so they can get their work done on time for result printing in the morning. What should the processor do?Correct Answer: Look up the laboratory's policy for handling such a situation and follow the policy.Discussion: The laboratory is not permitted to change a doctor's order in any way. By ordering the CBC the processor is ordering a test that the doctor did not specifically order and therefore makes the laboratory subject to a violation of the False Claims Act. By reviewing and following the laboratory policy the processor assures that the laboratory, the physician and the patient's best interests are met.

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Case Study 4

A busy laboratory is located in a 350 bed urban hospital that provides laboratory testing for the hospital and for the hospital's outreach testing program. A medical technologist in the microbiology department receives a call from a friend who works in a laboratory in a physician office. The physician is not a regular client of the laboratory currently but uses another laboratory for most of their work. The microbiologist knows that the sales department would like to get this account. The friend explains to her that she is doing a quality control check on her in-office microbiology testing and her regular laboratory will do it but is going to charge her for it. She asks the microbiologist if she will do it for free since it is quality control, not Medicare, and is not going to be billed to anyone.She tells the microbiologist that she would like to use the hospital lab for everything but her doctor insists on using the competitor. She indicates that the favor might help get the doctor to try the hospital laboratory for other tests. How should the microbiologist respond to this request?Correct Answer: Explain to her friend that if the hospital does the tests for no charge on the promise of other referrals, both the physician office and the hospital could be liable for violations of the antikickback statute.Discussion: The antikickback statute is implicated in this scenario because the free testing is solicited on the condition that other referrals may occur as a result of providing the favor. In fact, the solicitation itself is a violation of the law. The fact that Medicare patients are not specifically mentioned in the scenario is not sufficient to remove the risk. The technologist should also report the incident to the Compliance Officer and seek advise about what documentation, if any, should be kept concerning the incident.

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Case Study 1

A billing clerk is entering billing demographics on requisitions as a part of the normal days work. The department is under pressure to reduce accounts receivable, which means that the more clean claims that are filed, the better. This particular requisition is for a Medicare patient and has an LMRP test but does not have a diagnosis on it. She remembers that just a few requisitions before this one she had a requisition from the same doctor that had this same test on it that did have a diagnosis that allowed the test to be billed. She thumbs back in the pile and finds the previous requisition, notes the code that was used and adds it to the current requisition. This will help her meet the department goal of getting claims paid and reducing accounts receivable. Can she do this?Correct Answer: She should not do this because it is against the law to change diagnosis information on a requisition.Discussion: A laboratory employee should never change, add or use previously received diagnosis information for the purpose of making a test billable for the Medicare program or for any other insurance or payer. This is a form of fraud and for each claim submitted as a result of this activity, the laboratory is liable for a false claim and would have to pay the government back up to three times the reimbursement for the test and up to $10,000 for each claim submitted. If the employee is caught doing this, even if the employee is ignorant of the law and any laboratory policy prohibiting it, she must be disciplined along with her supervisor. Any employee who notices another employee doing this should correct the employee and report the incident to the department supervisor immediately.

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Advance Beneficiary Notice (ABN)

An Advance Beneficiary Notice (ABNs) is given to a Medicare beneficiary to inform him/her (or that person's representative) that Medicare may not provide coverage in a specific case (e.g., for a specific laboratory test). Entities who issue ABNs are known as “notifiers.” "Notifiers" can include physicians, practitioners, laboratories, and other suppliers of services that are paid under Medicare Part B. The "notifier" (e.g., the laboratory) must complete the ABN and deliver the notice to the affected beneficiary or his/her representative before providing the items or services that are the subject of the notice. The ABN must be verbally reviewed with the beneficiary or his/her representative and any questions raised during that review must be answered before it is signed. The ABN must be delivered far enough in advance that the beneficiary or representative has time to consider the options and make an informed choice. Once all blanks are completed and the form is signed, a copy is given to the beneficiary or representative. In all cases, the notifier must retain the original notice on file. Note: ABNs are never required in emergency or urgent care situations.

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Changes to ABN, Effective March 1, 2009

Beginning March 1, 2009, providers (including independent laboratories), physicians, practitioners, and suppliers are required to use a newly revised ABN form --Form CMS-R-131-- for all situations where Medicare payment is expected to be denied. The revised ABN replaces the existing General Use ABN, ABN-G (Form CMS-R-131G), and the Laboratory ABN, ABN-L (Form CMS-R-131L), and NEMB (Form CMS-20007). An example of the new form is included on this page as a resource. The revised form will continue to be used for notifying beneficiaries of Medicare denial reasons, but it may also be used to provide voluntary notification of financial liability. The revised ABN still permits pre-printing of lab-specific key information (blanks A - D) and still permits the use of the same denial reasons that were used with the former ABN-L (Blank E). Three commonly used reasons for noncoverage are: Medicare does not pay for this test for your condition. Medicare does not pay for this test as often as this (denied as too frequent). Medicare does not pay for experimental or research use tests. There must be at least one reason applicable to each item or service listed in Blank (D). The same reason for noncoverage may be applied to multiple items in Blank (D).

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HCPCS and CPT coding

The Healthcare Common Procedure Coding System (HCPCS) and the Current Procedural Terminology (CPT) codes are used to describe specific tests or services. The amount of payment for a test is dependent on the HCPCS or CPT code. HCPCS or CPT codes should be assigned under the supervision of the laboratory technical staff. Billing department employees should never change a HCPCS or CPT code without the approval of a manager or compliance officer. If a billing department clerk notices that a particular HCPCS or CPT code is being rejected by a payer they should report it to their manager. It is against the law to use the wrong HCPCS or CPT code for the purpose of causing or increasing payment for a test.

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ICD-9CM coding

ICD-9CM (International Classification of Disease, 9th Edition, Clinical Modification) codes are used for the classification of diseases and conditions, and for describing signs, symptoms and medical circumstances. These codes are used to indicate the medical necessity of a particular test. All employees who are directly or indirectly responsible for reporting to Medicare must be aware of these guidelines to prevent fraudulent claims: ICD-9 codes can only be supplied by the ordering physician or a representative of that physician. ICD-9 codes cannot be used from a previous laboratory order. If a physician supplies a narrative description instead of an ICD-9 code the laboratory must accurately translate that code using only certified coders. It is against the law to use the wrong ICD-9 code for the purpose of causing or increasing payment for a test.

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Local medical review policies (LMRPs)

Local Medical Review Policies (LMRPs) are published by Medicare for some laboratory tests. They are usually developed for tests that can be used for screening or diagnosis of disease. LMRPs use CPT codes to identify the tests and ICD-9 codes to determine when coverage is allowed. If an LMRP test is ordered by a physician, an ICD-9 code that is included in the LMRP must be given to the laboratory or the Medicare program will not pay for the test. It is against the law for a laboratory to change or add an ICD-9 code submitted by a physician. A laboratory should not submit a claim for an LMRP test that is not accompanied by an acceptable ICD-9 code. The Balanced Budget Act of 1997 made it illegal for physicians to order LMRP tests and not supply an ICD-9CM code with the order.

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Medical necessity

The Centers for Medicare and Medicaid Services (CMS), the US agency that administers the Medicare program, defines "medical necessity" as services or items reasonable and necessary for the diagnosis or treatment of illness or injury or to improve the functioning of a malformed body member. Medicare will not pay for any tests that CMS determines as unnecessary for diagnosis or treatment of disease.A laboratory may not submit a claim to Medicare or other government payers for any test it suspects is not medically necessary unless: The patient has signed an Advanced Beneficiary Notice, or A patient has requested the lab to submit such a claim for a determination by Medicare. Medicare does not pay for screening tests or tests that are ordered in the absence of signs or symptoms. Billing department employees are responsible for following all policies and procedures related to the submission of claims to reduce erroneous billings.

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Confidentiality

All employees have a responsibility to maintain the confidentiality of medical information. Medical information should never be discussed outside of the laboratory. Do not leave test orders or test results in areas where they can be viewed by patients. Do not discuss test results or any patient information in areas where patients can overhear the conversation. Be careful not to discuss confidential information on the telephone where patients can overhear the conversation. Employees should verify the identity of an individual requesting patient information.

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Inducements

It is against the law to offer or ask for money or favors to get a physician to order tests from a laboratory. This is known as an "inducement" or a "kickback." Laboratories should only give supplies to a physician for the drawing, processing, storing or transporting of specimens to the laboratory. The laboratory cannot provide supplies physicians use for their own purposes. The laboratory must monitor the amount of supplies provided to ensure that it matches the number of tests sent to the laboratory. The laboratory cannot give free tests except in the event of laboratory error. The laboratory cannot give free education to clients unless it is about the laboratory's services or policies. The laboratory cannot give excessive or expensive gifts or entertainment to physicians. The laboratory can give discounts, but the price must be above cost and at "fair market value."

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Phlebotomists and equipment in client offices

Laboratories may place phlebotomists or other employees in a physician office if all of the following are done: Employee only performs laboratory related tasks. There is a written understanding given to the physician about what the employee can and cannot do. Periodic audits are done to ensure the employee is following these policies. Laboratories may place printers, computers, fax machines or other equipment or products in client offices as long as they ensure that: The physician understands the equipment belongs to the laboratory. It is used for laboratory purposes like receiving reports or ordering tests. Periodic audits are done to ensure that the client is using the equipment only for laboratory related activities.

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Couriers and referral tests

The laboratory's couriers may not transport items except those related to the testing services offered by the laboratory. Couriers must follow all OSHA standards for the handling and transport of specimens. The laboratory is responsible for all tests it refers to other laboratories. Laboratory should not change CPT codes supplied by a reference laboratory without contacting the reference laboratory. The laboratory is responsible for all tests it bills to Medicare/Medicaid even if the test was performed by a reference laboratory.

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Marketing and record retention

The laboratory must ensure that its sales and marketing materials are clear and not deceptive: They should fully inform the physician about the appropriate use of laboratory tests and services. They should not be designed to induce unnecessary testing. The laboratory must have in place a document control system and record retention policy that ensures records are accessible in case of an audit or investigation. Records should be retrievable. Related documents should be linked.

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Utilization and other regulations

Laboratories must not induce physicians to order unnecessary tests through their marketing or education activities: They must monitor the use of laboratory services by their clients. They must correct any situation where something they did caused an unnecessary increase in test utilization. Cost Reports Hospital laboratories must ensure that information used in hospital Medicare cost reports is accurate and includes only those costs which are appropriate. Laboratories must follow all CLIA and OSHA regulations: failure to do so may result in a False Claims Act violation.

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Billing

Billing is the highest risk area for the laboratory because it generates the claims that are sent to Medicare and other government payers. Payment and payment errors are the focus of the OIG compliance guidance for the laboratory because of the revenue involved. Fraud and abuse is often perpetrated as a billing scheme. Nearly all laboratory functions can affect the billing of laboratory tests.

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Laboratory Billing Department Communication With Physicians and Patients

It is important that billing department employees are clear and accurate when communicating with physician office personnel and patients. Never guess at the answer to a question; ask if you are unsure. Do not speculate or express personal opinions. When requesting diagnosis information from the physician office staff, be careful to not lead them to give a billable code. The code must come from the patient's medical record. Billing department employees must accurately state laboratory policies and procedures, or forward the call to a supervisor to avoid misstatements and misunderstandings.

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Advance Beneficiary Notices (ABNs) and the Billing Department

A laboratory may not bill a Medicare beneficiary for a test unless it notifies the patient in writing before the testing is done that Medicare is not going to pay for the test. This notice is called an ABN. The ABN must contain the specific name of the test and give a specific reason the laboratory thinks payment for the test will be denied. The beneficiary should sign the ABN and a copy should be sent to the laboratory and one given to the beneficiary. The billing department must have evidence that the ABN has been signed before it bills a patient. A laboratory may bill Medicare even though it knows it will not be paid when it has evidence an ABN has been signed.

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Mycology: Hyaline and Dematiaceous Fungi
Match each of the names of the hyaline molds listed with the environmental conditions or natural objects with which it is most likely associated.View Page
The most helpful feature in differentiating the Zygomycetes from the other hyaline molds in the clinical mycology laboratory is:View Page
Saprophytic Cladosporium species may be difficult to differentiate from Cladosporium trichoides (Xylohypha bantianum) in culture as both produce chains of conidia separated by distinct scars or dysjuncters. Each of the following characteristics of Cladosporium trichoides are helpful in separating the two except:View Page
The disease with which the dematiaceous fungus illustrated in this photomicrograph is most likely associated is:View Page

Mycology: Yeasts and Dimorphic Pathogens
Match the names of the species of dimorphic fungi listed in the drop-down box with its corresponding yeast form as illustrated in the images.View Page
One of the characteristics common to the dimorphic molds is the ability to convert the mold forms to the yeast forms by incubating subcultures in enriched media at 35°-37°C. The upper image illustrates a subculture of a mold colony suspected of being a dimorphic fungus inoculated to the surface of blood agar and incubated for 3 days at 37°C. Note that the colonies have a prickly appearance, suggesting an intermediate stage of conversion. The lower image is a lactophenol blue mount of a portion of one of the prickly colonies. This fungus can be identified as:View Page
Each of the following dimorphic fungal infections have been observed in animals living in their natural environment except:View Page
Procedures for the rapid culture confirmation of suspected colonies of B. dermatitidis, C. immitis and H. capsulatum recovered from clinical specimens include:View Page
The colonies growing on the surface of this brain-heart infusion with blood agar plate were "converted" from a mold colony suspected of being Histoplasma capsulatum by incubating a subculture at 37°C for 5 days. The yeast forms that must be identified in mounts made from one of these colonies to confirm the identification are:View Page
Although care should be taken when working with all fungus cultures in the laboratory, personnel are particularly prone to develop laboratory acquired infections from the inhalation of airborne species of:View Page
Match each of the fungal species listed below with the appropriate category, indicating whether or not it has the capability of producing pseudohyphae on cornmeal agar.View Page
This photomicrograph is an acid-fast stained smear prepared from a yeast colony growing on ascospore agar. A helmet-shaped, red-staining, acid fast yeast cell is seen in the center of view at the tip of the arrow, against the background, blue-staining blastoconidia. The presumptive identification of Hansenula anomala was made. Predisposing conditions that may indicate that this isolate is more than a contaminant include:View Page

OSHA Chemical Hygiene (updated 2007)
Chemical Hygiene

In 1990 OSHA issued a Standard to replace Haz-Com specifically designed to: Meet the needs of laboratories with large varieties of chemicals. Mandate specific training for laboratory employees. This standard is called Chemical Hygiene (Standard # 1910.1450.)

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Right to Know

As an employee, you have the right to know the types of hazardous substances that are used in your workplace. Material Safety Data Sheets, or MSDS, are available in paper or electronic versions in each laboratory section and are there to inform you of these hazards. Ask your supervisor for the exact location.

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Fire and Explosion Hazard Data

Identifies any special precautions which should be taken during fire fighting procedures. This chemical is still flammable when diluted, and can be extinguished by an ABC fire extinguisher. Special fire fighting procedures included would not necessarily apply to a laboratory setting.

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Be Prepared!

Remember to read and be familiar with the MSDS and the Laboratory's Chemical Hygiene Plan before you start a job so that you will be prepared for any emergency.

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General Laboratory Precautions

Laboratory safety includes a number of precautions designed to protect you and your coworkers. Remember that: eating drinking smoking applying cosmetics or lip balm are forbidden in areas where chemicals are present.

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Personal Protective Equipment

Personal protective equipment is an essential way to protect yourself from the dangers of chemicals. You'll find on the label or MSDS exactly what kinds of clothing, gloves, and coverings you'll need to keep yourself safe. Also, the laboratory's chemical hygiene plan will include information about necessary personal protective equipment and engineering controls that will reduce your exposure to hazardous chemicals. At a minimum, safety goggles and rubber or nitrile gloves (not necessarily utility gloves) are necessary parts of your personal protective equipment.

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Fire Extinguishers

If there's a fire in the laboratory and chemicals are involved, there's a chance that some of the chemicals could react adversely with water. For that reason, the best fire extinguishers to use are ABC fire extinguishers. Using water to extinguish a chemical fire could actually fuel the blaze or cause chemicals to splatter.

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Monitoring

Certain chemicals in use in the laboratory, such as formaldehyde, are hazardous if your exposure to them is too prolonged. The amount of the chemical to which you can be exposed before possible danger is called the threshold limit value. Monitoring badges are used from time to time to measure your exposure. These are worn in the "breathing zone" for a certain period of time--often eight hours (for long-term exposure) or fifteen minutes (for short-term exposure). Based on the results of this monitoring, additional personal safety measures, such as ventilation or face-fitted masks, may be implemented for your protection.

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Common Sense Rules (continued)

Also remember to: Learn basic first aid measures. Read chemical labels. Read MSDS. Follow warnings and instructions. Use the correct protection. Practice sensible, safe work habits. Be knowledgeable about your laboratory's Chemical Hygiene Plan and the location in your laboratory of all reference materials on the hazards, safe handling, storage, and disposal of hazardous chemicals, including the location of Material Safety Data Sheets.

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Keep It Safe!

Your laboratory has provided you with training to protect yourself from chemical hazards in your daily work. But the only one who can keep you safe on the job every day is you. As a responsible member of the laboratory team, it is up to you to utilize safe work practices.

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OSHA Electrical Safety (updated 2007)
An adapter can be used to increase the number of available outlets in a laboratory area.View Page
Precautionary information that pertains to a laboratory instrument or appliance can ONLY be obtained by contacting the manufacturer.View Page
Introduction

Electrical hazards are present in all laboratories. Because of this, adherence to all electrical safety standards is essential in preventing electrical accidents.

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Ground

A ground is a conducting connection between an electrical circuit or equipment and the earth, or between an electrical circuit and some conducting body that serves in place of the earth.The purpose of a ground is to prevent the buildup of voltages that may result in a hazardous situation for the connected equipment and/or for the person operating the equipment.All electrical equipment in the laboratory that is not clearly marked as "double-insulated" must be grounded by using a three-pronged power cord.

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Space heaters

Electrical space heaters are prohibited unless they are approved for use and inspected by your facility's management department. Space heaters are not permitted in hospital sleeping areas or in laboratories containing flammable liquids or gases.

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Checks and Inspections

All laboratory instruments and appliances should be checked for ground integrity and current leakage before initial use, after repair or modification, and any time a problem is suspected.Periodic checks should be made on all electrical wires. If frayed wires are found, the equipment should be immediately removed from use and repaired.Report to your supervisor any shocks or tingling received from electrical equipment.

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OSHA Formaldehyde
Relevant OSHA Standards

1987 Haz-Com Standard is designed to help control employee exposure to chemicals on the job.1990 Chemical Hygiene Standard is specifically designed to meet the needs of laboratories with large varieties of chemicals, and to require specific training for laboratory employees.1992 Formaldehyde Standard is specifically for employees that work with formaldehyde. The goal was to reduce the risk of formaldehyde overexposure by establishing safe exposure limits.

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Where is Formaldehyde Used?

Formaldehyde is a preservative, ideally used in: Histology laboratories Autopsy laboratories Surgical pathology Anywhere biopsies are performed

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What is Formaldehyde?

Formaldehyde solution is a colorless, aqueous solution containing not less than 37% of formaldehyde or CH2O. It is usually supplied in 55 gallon drums.Formalin is a 10% formaldehyde solution that is commonly used in the laboratory.

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Material Safety Data Sheet

Formaldehyde has an MSDS (Material Safety Data Sheet).It can be found in your laboratory's MSDS book.Read the MSDS carefully!Ask questions.

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Your Laboratory

Your laboratory is committed to providing you with a safe working environment. It also expects you to do your part : Be a responsible member of the laboratory team. Use safe work practices.

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OSHA Formaldehyde (updated 2009)
Relevant OSHA Standards

1987 Haz-Com Standard (29 CFR 1910.1200) is designed to help control employee exposure to chemicals on the job. 1990 Chemical Hygiene Standard (29 CFR 1910.1450) is specifically designed to meet the needs of laboratories with large varieties of chemicals, and to require specific training for laboratory employees. 1992 Formaldehyde Standard (29-CFR 1910.1048) is designed specifically for employees who work with formaldehyde. The goal of this standard is to reduce the risk of formaldehyde overexposure by establishing safe exposure limits.

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Respirators

Respirators usually are not needed in a pathology laboratory. However, they are required if exposure exceeds the Action Level of 0.5 ppm. Respirators, if required, must be provided to you at no cost.

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In case of a spill...

Smalll spills (generally less than one liter) may in most instances by handled by laboratory or other employees. However, if you experience symptoms of over-exposure during the clean up, such as burning eyes, or throat irritation, immediately leave the cleanup area and get help from your institution's Spill Response team or other designated persons.Larger spills (generally greater than 1 liter) will usually require immediate assistance from the Spill Response team or other designees. There are several ways to clean up small spills, two of which are described below:1. Dike up the formaldehyde with absorbent pillows. Then dispose of these pillows in a sealed, formaldehyde-labeled container. 2. A chemical that reacts with and neutralizes formalin such as ALDEX® may be used to treat the spill.Your supervisor will show you the location of these emergency spill clean-up materials or discuss alternative procedures. Be sure to follow your own institution's policies and procedures in regard to formalin spills.

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What is Formaldehyde?

Formaldehyde solution is a colorless, aqueous liquid with a pungent odor. Concentrated formaldehyde solutions contain not less than 37% of formaldehyde or CH2O. These are usually supplied in 55 gallon drums. Ten percent (10%) aqueous formaldehyde solution, known as formalin , is almost universally used in the histology laboratory to fix and store pathology specimens, and, while still an important potential health hazard, is safer to use because of its lower formaldehyde concentration.

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Material Safety Data Sheet

The appropriate MSDS (Material Safety Data Sheet) for formaldehyde can be found in your laboratory's MSDS book or online. You should know where to access the MSDS and be familiar with its content. Direct any questions you may have to your supervisor.

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How is Exposure to Formaldehyde Monitored?

A monitoring badge is attached to your lab coat collar in the "breathing zone." for a specified length of time. This badge is then sent to an outside laboratory by your supervisor to determine your level of exposure to formaldehyde.

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Fume Hoods and other Controls

Engineering controls must be established to reduce formalin exposure to the lowest possible level. In most cases, chemical fume hoods or/and ventilated grossing stations serve as the primary engineering controls to reduce formaldehyde vapors. Rooms in which formalin is used may also require special direct exhaust ventilation. Formaldehyde should be dispensed or used in a chemical fume hood or other appropriately ventilated and approved work area. Check your laboratory's policies and procedures to be sure you use the engineering controls provided, as well as the required personal protective equipment.

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Packaging and Shipping Infectious Materials
What Records Must Be Maintained?

Training records must include: Employee name Most recent date trained Description of training Description, copy, or location of training materials Name and address of trainerThese records must be maintained throughout employment and 90 days thereafter, according to the US Department of Transportation (DOT). IATA and some laboratory regulatory agencies, including the College of American Pathologists (CAP), require repeat of training every two years. DOT requires training every three years. You will be able to print a certificate when you have completed this course that will certify your completion of training for packaging and shipping Division 6.2 (infectious) materials.

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Classifications of Hazardous Materials

The US Department of Transportation (DOT) classifies hazardous materials according to the risks that they pose. There are nine hazard classes: Class 1: Explosives Class 2: Gases Class 3: Flammable liquids Class 4: Flammable solids Class 5: Oxidizers/organic peroxides Class 6: Toxic and infectious substances Class 7: Radioactive material Class 8: Corrosives Class 9: Miscellaneous hazardous materials Within class 6 are two divisions: Division 6.1- poisonous material Division 6.2- infectious substanceA division 6.2 infectious substance is defined as a material known or reasonably expected to contain a pathogen. A pathogen is a microorganism or other agent (e.g., a prion) that can cause disease in humans or animals. The regulations that govern packaging and shipping a class 9, miscellaneous hazardous material, may also need to be reviewed by those who package and ship laboratory specimens. Dry ice is a class 9 hazardous material and, if used, requires special packaging, and specific labeling and marking on the outer package.

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Exempt Substances

Laboratory specimens that are unlikely to cause disease and do not meet the criteria for category A or B substances are not subject to Division 6.2 regulations. Specimens for which the hazardous materials regulation (HMR) does not apply include human or animal samples (including, but not limited to, secreta, excreta, blood and its components, tissue and tissue fluids, and body parts) being transported for routine testing not related to the diagnosis of an infectious disease. This includes specimens that are being sent for: drug or alcohol testing cholesterol testing blood glucose level testing prostate specific antibody (PSA) testing testing to monitor kidney or liver function pregnancy testing tests for diagnosis of non-infectious diseases such as cancer biopsies

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Exempt Specimen Packaging and Labeling

Although the specimens that were discussed on the previous page are not subject to the Division 6.2 requirements for packaging and labeling, it is important to remember that there are other packaging and labeling requirements that may apply, such as OSHA requirements included in the Bloodborne Pathogens Standard, healthcare facility requirements, and laboratory regulatory agency requirements, such as those of the Joint Commission and the College of American Pathologists. If the package is being sent by air, IATA, does require an exempt specimen to be marked as "Exempt Human Specimen," or "Exempt Animal Specimen." The DOT does not require this marking, so it would not be necessary if the specimen is being shipped by ground.

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Classification Scenario 1

A blood specimen is collected from a patient that is suspected of having Hepatitis B. The specimen will be sent via commercial carrier (e.g., Federal Express, DHL, or UPS) to a reference laboratory for further testing. What classification should be used for appropriate packaging and labeling? Work through the Classification Decision Tree.

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Classification Scenario 2

A blood specimen is collected from a patient suspected of having Hepatitis B. The specimen will be taken to the testing laboratory by the laboratory's own courier service using an exclusive use motor vehicle. What classification should be used for appropriate packaging and labeling?Work through the Classification Decision Tree.

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Classification Senario 3

A blood sample, collected from an outpatient, will be sent via FedEx to a reference laboratory for cholesterol screening. What classification should be used for appropriate packaging and labeling?Work through the Classification Decision Tree.

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Classification Scenario 4

A culture of Mycobacterium tuberculosis is to be sent by commercial carrier to the Public Health Laboratory. What classification should be used for appropriate packaging and labeling? Work through the Classification Decision Tree.

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Packaging Considerations

Several things need to be considered when you are determining how to package a laboratory specimen. These considerations include: Type of specimen Solid Liquid Classification Category A Category B Exempt Size of the specimen Temperature at which the specimen must be held during shipping Will dry ice be included in the package? The specimen components Does the specimen contain a preservative, such as formalin, that may be regulated? Mode of transportation Commercial ground Passenger air Cargo air Postal service Private or contract carrier using exclusive use motor vehicle

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Shipper's Declaration For Dangerous Goods- First Section

The first section of the form contains your laboratory's name and address (Shipper), and the name of the laboratory or organization where the specimen is being sent (Consignee). If an Air Waybill is used, its number should also be included on this form.

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Four scenarios will now be presented to evaluate your understanding of the material that has been presented on classifying, packaging, and labeling laboratory specimens for shipment. The scenarios are for your practice and will not be graded.Choose all the appropriate labels from the table below that must be used when packaging the substances described in the following scenarios:Scenario OneThree serum samples for hepatitis testing need to be sent via courier. The courier is an employee of your healthcare system. The specimens will be transported in a motor vehicle used exclusively for transporting specimens. What packaging labels are required? Choice Label Choice Label AGBH CIDJ EK FL View Page
A sputum specimen from a patient suspected of having tuberculosis is being sent by commercial ground carrier (FedEx, DHL, or UPS) to a reference laboratory. The package does not require a refrigerant.Which of these labels must be used on the outer packaging? Choice Label Choice Label AGBH CIDJ EK FL View Page
References

International Civil Aviation Organization. Technical instructions for the safe transport of dangerous goods by air. Doc 9284; 2005 - 2006 ed with amendment. National Laboratory Training Network. Packaging and shipping Division 6.2 materials. Georgia Public Health Laboratory; 2008. Sentinel Laboratory Guidelines for Suspected Agents of Bioterrorism. Available at: http://www.asm.org/ASM/files/LeftMarginHeaderList/DOWNLOADFILENAME/000000001202/Packing&Shipping11-18-05.pdf Accessed on February 13, 2009.US Department of Transportation Pipeline and Hazardous Materials Safety Administration. Transporting Infectious Substances Safely. Guide to changes effective October 1, 2006. Washington, DC; 2006.

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Parasitology Review
The avoidance of laboratory diagnosis techniques that utilize water is recommended for the identification of which of these parasites? View Page
Match each parasite listed here with the appropriate laboratory technique that may be used for its identification: Each answer may only be used once.View Page
The ideal stool sample for parasitic examination is one that is freshly collected and submitted to the laboratory at:View Page
Suppose that a stool sample was submitted to the laboratory for O & P examination. Macroscopic examination revealed a chalky-clay colored sample. If you were the alert clinical laboratory scientist on duty, which of the following would be the proper protocol would you follow in handling this sample?View Page
Suppose that a stool specimen was received in the laboratory for an O & P examination. The clinical laboratory scientist on duty performed direct wet preparations and found suspicious forms. An ethyl acetate concentration procedure was done, the top layer was examined, and no suspicious forms were seen. A slide of the sample was stained with Trichrome and again suspicious forms were noted. Which of the following is the most likely explanation for these discrepant results?View Page
A 44 year old female immigrant from Southeast Asia presented to the local clinic complaining of fever, chills, diarrhea and weakness. Patient history revealed that the woman worked in a research laboratory in her homeland. Her primary project was to develop an effective insecticide for the dreaded sandfly. The doctor decided to culture her blood for parasites. This form, measuring 14 µm, was recovered. The patient is most likely suffering from:View Page
A 65 year old Asian female presented to the emergency room exhibiting severe abdominal pain, fever and diarrhea. Examination revealed an enlarged liver that was tender to the touch. Patient history revealed that the woman worked in a fish processing plant for years prior to moving to the United States. Her diet was heavy in raw fish. Stool and duodenal contents were collected and sent to the laboratory for cultures and parasite examination. The cultures were unremarkable. This suspicious form was seen in both specimen types. It measures 27 µm by 14 µm. This patient is most likely suffering from:View Page
A fresh stool sample was submitted to the laboratory for parasitic examination on a 30 year old male who presented to a local clinic complaining of gastrointestinal discomfort and overall weakness. The only patient history available about the patient was that he was here job hunting and that he is originally from rural Mississippi. The sample was immediately processed and this suspicious form was seen. No other suspicious forms resembling eggs were seen. The patient is most likely infected with:View Page
A stool collected at a local doctor's office was received in the laboratory for parasitic examination. The sample was not received in fixative and due to the new courier system did not arrive in the lab within the traditionally acceptable time frame. Due to logistical difficulties of the patient collecting and submitting another sample, the laboratory director authorized the sample to be processed. The comment "specimen delayed in transit, please evaluate results accordingly" was included in the report. These two suspicious forms were seen upon examination of the specimen. Label these two forms:View Page
A stool was received in the laboratory for parasitic examination on a 49 year old female who just returned from missionary work in numerous third world countries around the world. The patient had been suffering from mild diarrhea over the past two weeks. These two suspicious forms were seen. Form 1 measures a mere 6 µm whereas form 2 measures 35 µm. Label these two forms:View Page

Pharmacology in the Clinical Lab: Therapeutic Drug Monitoring and Pharmacogenomics
Introduction

Therapeutic drug monitoring and pharmacogenomics are both pharmacy-related areas within the clinical laboratory. Although each is considered a sub-discipline within laboratory medicine, the two fields overlap significantly. In this course, we will provide an overview of each of these laboratory sub-disciplines and discuss the utility, rationale, and practice of each one.

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Laboratory Methods

Immunoassay is the most common technique used by clinical laboratories for therapeutic drug monitoring. Antibodies that recognize drugs can be developed. Although most drugs are much too small to evoke an immune response, scientists can conjugate drugs to immunogenic proteins to produce antibodies that recognize drug-specific epitopes. There are several methods that utilize the principals of immunoassay for detection and quantification of therapeutic drugs in serum. Some of these methods are: Particle-enhanced turbidimetric inhibition immunoassay (PETINIA) Fluorescence Polarization Immunoassay (FPIA) Chemiluminescent assays

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Clinical Utility

The ultimate goal in measuring CYP450 function or identifying polymorphisms is to predict effective therapeutic doses and responses in patients.Polymorphisms are identified using molecular techniques (allele-specific PCR, restriction digests, sequencing, hybridization assays, bead-based systems, microarrays, pyrosequencing, et al).Although most clinical labs do not offer PGx testing, reference labs are beginning to market these tests. For example, one reference laboratory in the Midwest that offers CYP2D6 profiling measures about one dozen of the most common and significant mutation sites on this enzyme. This allows for detection of approximately 98% of the known CYP2D6 polymorphisms. The laboratory then generates a report which will advise the physician on the patient's drug-metabolizing status.Estimates show that 6-10% of the general population have a complete deficiency of CYP2D6, with the prevalence of mutations varying from <1% to as much as 21% within a given population.

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Genotype versus Phenotype

Phenotyping involves measuring the metabolism of a probe drug. For example, with CYP2D6, dextromethorphan or debrisoquine can be given to a patient to see how well the drug is metabolized. Both these drugs are safe and extensively metabolized by CYP2D6. By measuring the parent drug and the metabolite in urine, the metabolic capacity of a CYP450 enzyme can be estimated. Such testing is complex and tedious, however, and has not become routine in clinical laboratories. Therefore, genotyping is likely to be the main tool that is used for assessing the PGx of a patient.

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Phlebotomy
Case

James Brown, a phlebotomist from the laboratory went to the second floor of Memorial Hospital to draw a STAT BMP (chem-8), CBC, and PT on a patient. The patient was in critical condition so the lab results were crucial for treatment. James quickened his pace in order to speed up the result time. He collected the specimens and took them back to the lab. However, the technologist in hematology and coagulation notified him that he would need to recollect the specimen because the CBC and PT were clotted.

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Case

A phlebotomist from the laboratory at Midtown Memorial Hospital was working evening shift. Her shift ended at 11 PM and it was 10:30 PM. She suddenly got orders for a STAT blood culture on the second floor. The order specified blood culture times two, 30 minutes apart. The phlebotomist went to the patient’s room and decided to collect both blood cultures at the same time form the same site so she would be able to leave on time without having to come back in thirty minutes to collect the second set. She also wanted to “save” the patient from an extra stick. While the phlebotomist was preparing for the collection, she realized she didn’t have any Betadine on her tray, and decided she would just clean the site twice with alcohol. She finished the blood culture collections and was able to leave by 11 PM.

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Unsatisfactory specimens

Poor and unsatisfactory specimens pose significant problems : They can cause misleading laboratory results.Unsatisfactory specimens must be rejected by the laboratory. The patient must then undergo another venipuncture to get a better specimen. It costs time & money to redraw the specimen.The credibility of the laboratory is reduced if too many unsatisfactory specimens are drawn.

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Labeling errors

Labeling errors are the most common cause of incorrect laboratory results.If detected, the incorrectly labeled specimen will be rejected.If undetected, it will produce incorrect results which might adversely affect your patient’s care. 

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What is a phlebotomist’s role in a health care facility?

The phlebotomist collects blood & other specimens which ultimately provide doctors and nurses with laboratory test information critical to patient care.He or she therefore plays a vital role in any health care system.

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Laboratory work-flow cycle

The work flow of any medical laboratory involves these basic steps: Physician orders lab tests. Order is received in lab. Work list and labels generated by lab. Phlebotomist is dispatched to patient.

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Work-flow cycle: patient ID to specimen processing

Phlebotomist positively identifies patient. Phlebotomist draws and labels blood specimen. Specimen is transported to laboratory. Specimen is accessioned and processed in lab.

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Work-flow cycle: test performance to treatment

Laboratory performs analytical tests. Lab results are returned to physician. Physician treats patient based on results of lab tests.

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Laboratory work-flow cycle: phlebotomist role

As a professional phlebotomist, you have a critical role in this basic work-flow cycle. The rest of this program contains the information you need to begin training in this important profession.

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Quality Control
What is Quality Control?

Quality control (QC) is a system used to maintain a determined level of accuracy and precision. Proper quality control helps ensure that reported results of patient laboratory testing are correct. Quality control applies not only to specimen testing, but also to collection, storage, and transportation.

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Assayed and Unassayed Controls

Assayed controls have been analyzed by the manufacturer so that the range of values for the analytes they contain is known. Unassayed controls are unknowns. The laboratory purchasing the controls must determine the concentration of each analyte.

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Calibration Curve

Standards are also used to determine the linearity of the testing instrument. This is done by plotting a calibration or standard curve. Most testing instruments must be operated within a linear range. The Clinical and Laboratory Standards Institute or NCCLS defines linearity as “the measure of the degree to which a curve approximates a straight line.The examples to the right show linearity because a change along the x-axis shows a corresponding change along the y-axis, whether the x-value is low or high.

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External Quality Control (1)

External quality control (also called proficiency testing or PT) evaluates a laboratory’s testing results by comparing them to those of similar laboratories. Specially prepared specimens are obtained by multiple laboratories from proficiency testing program sponsored by professional societies, such as the College of American Pathologists or the American Association of Bioanalysts. These "unknowns" are then tested by multiple laboratories, and results are returned to the proficiency testing program. For external quality control, the laboratory must use its routine testing methods.

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External Quality Control (2)

Validated material for the different testing areas in a laboratory (such as chemistry, hematology, and microbiology) are provided several times a year. Participating laboratories test the specimens and return results to the proficiency testing source. The laboratory’s performance is then evaluated using the comparative method mean as the target value plus or minus a defined limit. In general, the evaluation report will show: number of laboratories comprising the peer groupcomparative mean of the group for that particular analyte the laboratory’s performance compared to the peer group whether the performance was satisfactory or unsatisfactory.

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The Levey-Jennings Chart's Inventors

Daily Documentation and evaluation of quality control is vital to diligently monitor sources of error. One of the most commonly used methods for documentation is the Levey-Jennings control chart (often referred to as the L-J chart). In 1931, Dr. Walter Shewhart, a scientist at the Bell Telephone Laboratories, proposed applying statistical based control charts to interpret industrial manufacturing processes. In 1950, S. Levey and E.R. Jennings suggested the use of Dr. Shewhart’s control chart in the clinical laboratory.

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CUSUM Example: Plotting Control Data

To illustrate the use of CUSUM in the laboratory, we'll use daily control values for glucose testing. First, we'll list daily control values under "daily results." Then, we'll calculate mean by using formula A. Next, we can find the difference from the mean for each result, and square that result for the two relevant columns. Using all of the squared differences from the mean, we can find the standard deviation using formula B. Using the mean from formula A and the standard deviation calculations from formulas B and C, we can plot our data points on the Levey-Jennings chart. Formula D helps us calculate the coefficient of variation (CV), which expresses SD as a percentage of mean value and is more reliable for comparing precision at different concentration levels. The lower the CV the greater the precision.

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Verification of Performance Specifications for Nonwaived Testing

On April 24, 2003, the Clinical Laboratory Improvement Amendments (CLIA) Final Rules went into effect.As of that date, each laboratory that introduces a nonwaived, unmodified, FDA-cleared or approved test system must do the following before reporting patient test results: Demonstrate that it can obtain performance specifications comparable to those established by the manufacturer for the following performance characteristics: Accuracy. Precision. Reportable range of test results for the test system. Verify that the manufacturer's reference intervals (normal values) are appropriate for the laboratory's patient population.

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Verification for Nonwaived Testing

Laboratories that do nonwaived testing must verify that they can obtain performance specifications comparable to those established by the manufacturer. Generally this can be accomplished by doing split sample comparison studies with another laboratory to estimate any inaccuracy or bias, plus linearity studies to estimate imprecision and determine the reportable range. The laboratory can do studies to determine its own reference ranges or the laboratory director can document that the manufacturer’s ranges are appropriate. For those laboratories doing nonwaived tests that have been modified or developed in-house, additional verification studies are required.

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A laboratory developing a new testing procedure runs the test on 1,000 patients. Later confirmation shows that the experimental procedure produced remarkably few false negatives, but many false positives. The experiment test is therefore:View Page
What is internal quality control?View Page
Match the type of quality control to the benefits it provides.View Page
A Quality Control Exercise

For this problem, you may need to work off-line. After you have evaluated the data on the following page, return to the course and answer the accompanying questions. Problem You are the only full-time employee at a small clinic's laboratory. You use an assayed control for your glucose determinations. The manufacturer's printed values for the present lot number are: Level 1 Control Mean: 72 mg/dL Standard deviation: +/- 2 mg/dL Level 2 Control Mean: 281 mg/dL Standard deviation: +/- 12 mg/dL The table on the next page shows the control results for the first twelve days of testing for the month. Plot your QC results on a Levey-Jennings chart and evaluate your data.

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Red Cell Disorders: Peripheral Blood Clues to Nonneoplastic Conditions
G6PD deficiency

A ten-year-old boy came to a physician's attention because of recent jaundice and icteric sclerae. The immediate laboratory work revealed: Hct 24%(normal 36%-47%), MCV 79.5 fl (normal 78-95fl),RDW 13%(normal 11.5-15.0%). His blood smear findings are reflected in these photomicrographs. Note particularly the spherocytes in the upper picture. Some resemble a half-blister with the other half of the cell containing solidly-staining hemoglobin. These are called eccentrocytes. When present, they should trigger a search for red cell hereditary G-6PD deficiency and the oxidant that triggered hemolysis. These morphological findings are only clues; specific testing for G-6PD deficiency should be performed. The blue arrows in the upper photomicrograph are directed toward solid-staining spherocytes in which the cell membrane is beaded by inclusions wrapped within the cell membrane, suggesting the remains of denatured hemoglobin. Included on the smear is a target cell, several acanthocytes, a smudge cell, and a few schistocytes. The lower photomicrograph is supravital staining of affected red blood cells, verifying the presence of Heinz bodies. This disorder was first recognized during the Korean war in 10% of black American soldiers given the antimalarial drug primiquine.

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Reticulocyte identification

Reticulocytes are red blood cells prematurely released from the bone marrow. On a Wright-Giemsa stained blood smear, they appear as polychromatic macrocytes. Their presence in the peripheral blood may suggest hemolysis or bleeding. Their presence is expressed as a percentage of the red cell count: newly born= 3-7%; up to one week of age=1-3%; >one week =0.3-1.8%. Automated or manual methods may be used to enumerate reticulocytes. In clinical context, retics must be separated from debris, precipated stain, Pappenheimer bodies, Howell-Jolly bodies, and Heinz bodies.

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Dimorphic RBC population

Illustrated in the photomicrograph of a peripheral smear are two populations of erythrocytes. Approximately 50% of the erythrocytes are normal size and contain a full complement of hemoglobin. The patient had received blood transfusions. The transfused red blood cells are the normocytic, normochromic red cells. Admixed are microcytic erythrocytes and larger erythrocytes, some faintly mottled or smudged, suggestive of reticulocytes. This picture represents a hemolytic process with a reticulocyte response. A similar dimorphic red cell population appears following erythropoietin therapy. It is important to recognize when a population of cells in the peripheral smear is not in context with anticipated laboratory findings and the clinical situation.

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References

Glassy, Eric F.,(Ed). Color Atlas of Hematology: An Illustrated Field Guide Based on Proficiency Testing. 1998. College of American Pathologists Hematology and Cliical Microbiology Research Committee. College of American Pathologists, Northfield, IL 60093-2750.Hookey,L., Dexter, D., Lee,D. H. The Use and Interpretation Of Quantative Terminology In Reporting Red Blood Cell Morphology. Laboratory Hematology 7:85-88, 2001.Peterson P, Blomberg DJ, Rabinovitch A, Cornbleet PJ. Physician Review of the Peripheral Blood Smear: When and Why. For the Hematology and Clinical Microscopy Resource Committee of the College of American Pathologists. Laboratory Hematology 7:175-179, 2001

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Reporting of laboratory data in regard to blood cell abnormalities

Laboratory data must be presented to clinicians in a user friendly way to promote effective decision making. Databases must be designed to provide clear information that leads quickly to the best patient care outcome. We continue learning how to collect and retrieve laboratory data from our machines, but we are not always in tune to how entry and retrieval of data is geared to and, more directly, influences patient care outcomes. Examples of blood cell abnormalities on a peripheral blood smear that may immediately direct the physician to a specific diagnosis are: (1) presence of target cells as found in thalassemia or hemoglobinopathies and target cells in liver disease, particularly with obstructive jaundice; (2) burr cells as a signal of chronic renal disease and uremia; and (3)atypical neutrophil inclusions relating to genetic disorders. Critical appraisal of such observations could add valuable clues for a diagnosis. Laboratory professionals must establish a set of principles for orderly observation of blood cell morphology, have a clear vision of the applications of their work, and understand the potential clinical implications of their reports and interpretations. Emphasis on values and relevance focuses on patient care outcomes and their dependency on prompt availability of results and contextual interpretations.

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Criteria for peripheral blood smear review

Initial analysis of the peripheral blood picture is made in most clinical laboratories with an automated instrument. Samples are selected for further analysis when quantitative or qualitative abnormalities beyond a defined standard are found. The following are examples of quantitative RBC abnormalities that may prompt a blood smear review. Each laboratory, however, should develop its own guidelines: Hgb: < 8 or >18 g/dL (<10 or > 21g/dL in a newborn)Hct: <20% or > 60% in adults (<40% or >65% in a newborn)MCHC: <29 g/dLMCV: <69 femtoliters (fl) or >110flFlags generated by the hematology analyzer that indicate possible red cell abnormalities or spurious resultsAny of these findings should be followed up with a peripheral blood smear review.

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Guidelines for standard reports

In a study on the reporting of red blood cell morphology abnormalities conducted in Ontario, Canada (Hookey L, Dexter D, Lee DH, Laboratory Hematology 7:83-88, 2001), fewer than 50% of 33 participants used the same term to describe the quantitative frequency of peripheral blood abnormalities. Seven blood smears, each containing one of several abnormal erythrocytes-- schistocytes, teardrop cells, acanthocytes, and Howell-Jolly bodies--were evaluated by 32 participants. The participants were asked to document their evaluations from a list of quantitative terms. There was a heterogeneity in the use of terms "rare," "slight," "occasional," "few," "mild", "present," "moderate," "many," and "marked." Choices of terms were subjective without points of reference. Guidelines for establishing standardized qualitative estimations of abnormal erythrocytes in the peripheral smear are presented as follows: 1+ = 2 - 4/Oil Immersion Field (OIF) 2+ = 5 - 7/OIF 3+ = 8 - 10/OIF 4+ = >10/OIF. The terms "few," "moderate," "many," and "marked" may be substituted for the 1+ - 4+ grading system, but only when their specific points of reference are universally understood in tandem with the above guidelines. A comment should be triggered if any erythrocyte abnormalities are seen in numbers >3/OIF including, but not limited to, polychromasia, basophilic stippling, nucleated RBC's, and Howell-Jolly bodies. Rouleaux or RBC agglutination are important findings and must be documented.

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Qualitative reports: Follow-up

Any review of a peripheral smear is highly subjective; therefore, each laboratory must establish its own guidelines for quantitating observations and issuing reports in a consistent format. The key question for the laboratory is "How will the clinician use the terms of qualitative results in the reports issued to decide on the next course of action with this patient?" Formats for reporting have been geared more toward the needs of instrumentation facilitation and computer management than toward needs of access and understanding by clinicians working to improve patient care outcomes. Evidence based medicine (EBM) is the formal term used for the process by which research evidence, collective clinical experience, and the user friendly rendering of testing results are integrated to evaluate patient care outcomes.

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After a review of the peripheral smear represented by this photomicrograph, which report is the most appropriate for documentation of the findings?View Page

Red Cell Morphology
Summary

It is important to differentiate in vitro changes which are secondary to preparing the slide, from in vivo morphology, which is the result of the pathophysiological condition of the patient. Examining erythrocytes in the critical viewing area is extremely important in making this distinction. The determination of the clinical significance of the morphology reported is the responsibility of the physician, who must correlate the blood smear findings with the clinical diagnosis, and other laboratory parameters.

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More Acanthocytes

Acanthocytes can also be seen in this slide. Alcoholic cirrhosis is the most common source of acanthocytes seen in blood smears in the laboratory (10-50%). Other sources are lipid disorders and a small number following splenectomy.

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Routine Venipuncture
What is Venipuncture?

Venipuncture is the collection of blood from a vein. The person having the responsibility for the performance of the venipuncture may be a phlebotomist who is a part of the laboratory staff, or he/she may be another healthcare professional that has been trained to perform this duty. In this course, we will refer to the person performing the venipuncture as the phlebotomist.

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Tools of the Trade

A variety of tools are available to ensure a safe and successful venipuncture. It is important to know which tools to choose and how to use them correctly so that an adequate specimen is collected for laboratory testing.

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What is a Hidden Error?

Hidden errors are those that cannot be detected or corrected by the laboratory analyst prior to testing. Most often these errors can be prevented by the phlebotomist following correct venipuncture procedure for every procedure, every time.Hidden errors include hemoconcentration, incorrect order of draw, and (the most serious of all errors) misidentification of patient or specimens. Because these errors often are unknown, the analyst may inadvertently report erroneous patient results which could be harmful to the safety and well-being of the patient. Condition What is it? How does it happen? What is the Result? Hemoconcentration Blood pools at site of venipuncture Tourniquet is applied for a prolonged period of time Test results may be inaccurate because blood components move between blood and tissues Pouring Blood between tubes Mixing contents of two or more tubes Removing top of tube to combine contents of one tube with another Inaccurate test results due to over or under dilution or incorrect anticoagulant Clots form due to lack of mixing Patient may have to be redrawn Incorrect patient identification and incorrect specimen labeling Using the wrong name to label a specimen Failure to positively identify EVERY patient using 2 unique identifiers BEFORE beginning venipuncture Failure to label EVERY specimen in the presence of the patient Failure to concentrate fully on the task Results reported to caregiver for wrong patient Compromises patient care; may be life-threatening

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Labeling Specimens

All specimens must be labeled in the presence of the patient at the time of collection. Inaccurate or incomplete labeling may result in rejection of the specimen by the laboratory. Unlabeled specimens will automatically be rejected by the laboratory. When labeling a specimen for the laboratory, the following information must be included: Patient's first name and last name Hospital medical record number, date of birth or alternate unique patient number Collector's ID Time the specimen was collected Date the specimen was collectedA phlebotomist must NEVER pre-label specimen containers. This can result in specimen mix-up and potentially disastrous consequences for the patient.

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Correct Fill

Fill blood collection tubes completely (until vacuum is exhausted) to ensure the correct blood to anticoagulant ratio necessary for accurate patient results. Specimens may be rejected by the laboratory if the tube is short-filled or over-filled. To avoid short-filling of tubes, the phlebotomist must ensure that the blood flow stops completely before removing the tube from the needle. When using a winged device (butterfly) to collect blood for coagulation studies (e.g., protime, aPTT), the phlebotomist must draw a light blue top "waste" tube before attaching another light blue top tube for testing. If the air in the tubing of the winged device is not displaced into a waste tube and is drawn into the tube used for testing, the tube used for testing will short-fill. The laboratory may reject the specimen because of invalid blood to anticoagulant ratio.

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Order of Draw

Blood collection tubes must be filled in a specific order to avoid specimen contamination from the additive in the preceding tube. The following order of draw is an accepted laboratory standard. 1. Tubes or bottles for blood cultures 2. Light-blue top tubes (sodium citrate) 3. Serum tubes (with or without clot activator) 4. Green top tubes (sodium or lithium heparin) 5. Lavender or pink top tubes (Potassium EDTA) 6. Gray (Sodium fluoride and sodium or potassium oxalate)

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Pre-analytic and hidden errors can greatly affect a laboratory result.Match the error listed below with the cause from the drop-down box.View Page
What are Pre-analytical Errors?

Pre-analytical errors are errors that occur prior to the testing process. Hemolyzed specimens, clotted specimens, incorrect tube type, and inadequate tube fill can all produce pre-analytical errors. Fortunately, many of these errors can be detected by the laboratory analyst so that corrections can be made before testing begins or before resulting and reporting the test. Unfortunately, the correction that needs to be made usually involves redrawing the patient. The table on the following page lists several preanalytical errors that can occur during the phlebotomy procedure.

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References

Clinical and Laboratory Standards Institute (CLSI). Collection, Transport, and Processing of Blood Specimens for Testing Plasma-Based Coagulation Assays; Approved Guideline. Fourth ed. CLSI document H21-A4. NCCLS. Wayne, PA: 2003.Clinical and Laboratory Standards Institute (CLSI). Procedures for the Collection of Diagnostic Blood Specimens by Venipuncture; Approved Standard. Sixth ed. CLSI document H3-A6. NCCLS. Wayne, PA: 2007.Clinical and Laboratory Standards Institute (CLSI). Procedures for the Handling and Processing of Blood Specimens; Approved Guideline. Third Edition. CLSI document H18-A3. NCCLS. Wayne, PA: 2004.Ernst DJ. Applied Phlebotomy. Baltimore, MD: Lippincott Williams & Wilkins: 2005.Lowe B. Reinforcing safety sticklers. Advance for Medical Laboratory Professionals. May 2004; 16:2A-3A.The Joint Commission. Patient Safety-2009 National Patient Safety Goals. Available at: http://www.jointcommission.org/PatientSafety/NationalPatientSafetyGoals/. Accessed July 18, 2009.

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Blood Collection Tubes

Most blood collection tubes contain an additive that either accelerates clotting of the blood (clot activator) or prevents the blood from clotting (anticoagulant). A tube that contains a clot activator will produce a serum sample when the blood is separated by centrifugation and a tube that contains an anticoagulant will produce a plasma sample after centrifugation. Some tests require the use of serum, some require plasma, and other tests require anticoagulated whole blood. The table below lists the most commonly used blood collection tubes. Tube cap color Additive Function of Additive Common laboratory tests Light-blue 3.2% Sodium citrate Prevents blood from clotting by binding calcium Coagulation Red or gold (mottled or "tiger" top used with some tubes is not shown) Serum tube with or without clot activator or gel Clot activator promotes blood clotting with glass or silica particles. Gel separates serum from cells. Chemistry, serology, immunology Green Sodium or lithium heparin with or without gel Prevents clotting by inhibiting thrombin and thromboplastin Stat and routine chemistry Lavender or pink Potassium EDTA Prevents clotting by binding calcium Hematology and blood bank Gray Sodium fluoride, and sodium or potassium oxalate Fluoride inhibits glycolysis, and oxalate prevents clotting by precipitating calcium. Glucose (especially when testing will be delayed), blood alcohol, lactic acid

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Scenario Conclusion

When the results on Mr. John Ready were called to the nurse, she was very surprised that the result of his CBC was normal. The nurse explained to the laboratory technologist that Mr. John Ready had a known diagnosis of lower GI bleeding. His hemoglobin had been very low for the past 24 hours because of the internal bleeding, and she thought it was very surprising that his hemoglobin had normalized so quickly without having received a blood transfusion. Mr. Ready’s doctor decided the patient should be redrawn to ensure a correct result. The nurse further questioned if the phlebotomist could possibly have drawn the wrong patient because earlier that day Mr. Ready had been moved to room 831, and room 825 was presently occupied by a patient named Walter Redding. If Julie had properly identified the patient by asking him to state his name and then checking the name and identification number on the wristband, she would have realized that the patient in 825 was the wrong patient.

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Semen Analysis
A semen specimen was collected three hours before it was brought to the laboratory for examination. What course of action should be taken?View Page
Materials Needed

The following is a list of materials needed for semen analysis. Laboratories will differ slightly in the equipment used. Use of this equipment will be described further in the later pages of this course. Materials needed include:graduated test tube or serological pipets with safety bulb to measure volumepH paper in neutral to basic range (e.g. 7.2-8.8)counting chamber and/or automated counting machineglass slides and coverslips for wet mount if motility and sperm count are to be assessed separatelyhand counterif dilution is donediluting fluid calibrated automatic pipetspositive pressure pipets and glass boreslight microscope with phase contrast objectives for sperm count and bright field objectives for morphology assessmentglass slides and fixative for morphology slidesset-up for performing Papanicolaou or other morphology stainingEvery laboratory should also have a copy of the "WHO Laboratory Manual for the Examination of Human Semen and Sperm-Cervical Mucus Interaction", published on behalf of the WHO by Cambridge University Press. The fourth edition was published in 1999.

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White cells in semen

Round cells in semen are of two types: immature sperm and white blood cells. To determine the percentage of white blood cells (specifically granulocytes) a special leukocyte screening test must be done. This test involves staining for the peroxidase enzyme present in the granulocytes.The 1999 WHO manual contains a protocol for doing this test (Appendix III). There is also at least one test kit on the market for this assessment (Leukoscreen: Bioscreen, Inc.).Laboratories with particular expertise in doing CBC and assessing granulocytes in stained blood smears may be able to do a differential count by this method rather than using a biochemical test for leukocyte screening.

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Prerequisites

The basic laboratory skills that you will need to do a semen analysis include: Using a microscopePerforming manual cell counts and doing calculations to determine the concentration of those cells per milliliter of fluidMeasuring volumeMeasuring pHMeasuring viabilityKnowledge of OSHA regulations for handling potentially infectious human fluids

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Morphology: What this presentation will teach you.

This presentation will describe basic aspects of WHO III assessment. For information on details of some of the morphological assessments you will need to review information presented in the 1992 WHO III manual.Learning to do strict morphology assessments is more complicated than learning WHO III and generally requires that the technician take one of the many hands-on laboratory courses offered periodically around the country. Details of strict morphology assessment are presented in the 1999 WHO IV manual.

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Summary: Reference values

The following are reference values for a normal semen analysis. It should be noted that these are recommended reference ranges only and that they may require adjustment for your particular laboratory or region of the country:Liquefaction: ≤30 minutesVolume: ≥2.0 mlColor: white, yellowish, grayViscosity: non-viscouspH: ≥7.0Sperm count: ≥20 million / mlMotility: ≥50%Leukocytes: ≤1 million / mlWHO III Morphology: ≥30%Strict Morphology: ≥14% In addition some people find it useful to have a total motile count (TMC). This is calculated by multiplying the concentration x the percent motility x the volume. Normal TMC is 10 million or greater.

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Viscosity

Most semen is somewhat viscous. Liquefaction should be complete before viscosity is assessed. Semen viscosity can be determined by trying to draw the specimen into a wide bore pipette. Normal semen can be dropped from a pipette in single droplets. Some laboratories report viscosity on a scale from 0-4. Others report the results as "non-viscous", "slightly viscous", "very viscous" and so forth. A specimen that is more viscous than normal after liquefaction may have reduced sperm motility. During sexual intercourse, hyperviscosity can prevent the sperm from reaching the cervix.

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Viability

Viability is a measure of the percentage of cells that are alive. Since motile cells are inherently viable a viability assessment may not be necessary when motility is high. Most laboratories set a minimum motility after which a viability will also be performed. To assess viability, place a drop of semen on a slide. Add an equal volume of a vital stain such as trypan blue. Cover with a coverslip. Allow color to develop for several minutes, but not more than 5 minutes. Count 100 cells (both motile and non-motile cells) on each of 2 slides. During the count differentiate between the white cells (living) and blue cells (non-living).

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Collection (continued)

Other aspects of specimen collection that must be considered are the temperature of the specimen and the time needed to transport it to the laboratory.Ideally, the specimen should be collected in a room at the testing site.If on-site collection is not possible, the specimen should be kept at body temperature (37°C) from the time of collection until it arrives at the laboratory. This can be facilitated by holding the container close to the body, for example by carrying it in an inside pocket.Semen should arrive at the laboratory as soon as possible after collection, preferably within one hour.Lubricants should not be used for collection unless absolutely necessary as most lubricants are toxic to sperm. If lubricant must be used then non-toxic forms such as KY jelly or cooking oil should be the only options.

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Collection

Accurate semen analysis results require appropriate sample collection. Patients must receive detailed directions for proper specimen collection and transport. Directions should be in writing. Specific instructions should include: The period of abstinence prior to collection should be between 2 and 5 days.The entire specimen must be collected because the different portions have varying concentrations of spermatozoa.An appropriate collection container must be used.Each laboratory should designate an appropriate, wide mouth, collection container.Each lot of collection containers should be tested to ensure that it is non-toxic to sperm.Alternative collection containers should be discouraged because their level of toxicity is unknown.Use of condoms for collection should be discouraged particularly when the purpose of the semen analysis is to test for fertility. Some condoms are toxic to sperm. Collection in condoms often results in inaccurate results for semen volume and other parameters.

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Microscopic Examination of Semen

Microscopic examination of semen includes assessment of: sperm concentration percent motility percent viability cellular elements other than spermatozoa sperm morphology Sperm counting will be covered in this section. Assessment of other cellular components and of morphology will be covered in the next sections. For assessing count, motility, viability and other cellular components your laboratory will require a phase contrast microscope with 10x oculars and objectives up to 20x. For assessing morphology you will need bright field objectives up to 40x and 100x (oil immersion).

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Sperm counting methods

Sperm can be counted either manually or by automated methods. Although automated counting has some advantages for assessment of motility parameters, manual counting is still performed by most laboratories. There are several manual counting methods available for semen. These include:Neubauer hemacytometerMakler chamberCellVu (Millennium Sciences, Inc)MicroCell (Conception Technologies) The Makler, CellVu, and MicroCell methods have the advantage of requiring no dilution of the semen. Since semen is viscous, accurate dilution can be problematic. These methods also allow counting of motile and non-motile sperm at the same time and thus avoid the need for separate assessment via wet mount. Each laboratory should determine the best most reproducible method for their own situation, equipment, and expertise.

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Motility

Motility of a normal semen sample is 50% or greater.Sperm motility is important because sperm must be moving in order to penetrate the cervical mucus, travel to the fallopian tube, and fertilize ova.Accurate motility evaluation requires that the temperature be standardized. Some laboratories read motility at 37°C while others routinely report motility at room temperature. The temperature of the assessment should be specified in the final report.

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Requirements for the microscopic examination of semen

For assessing count, motility, viability and other cellular components your laboratory will require a microscope with 10x oculars and phase contrast objectives up to 20x. You will also need hand counters.For assessing morphology you will need bright field objectives of 40x and/or 100x (oil immersion).You will also need counting chambers, glass slides and coverslips and a method for staining sperm for morphology assessment.

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Assessing sperm motion parameters

In addition to determining the percent motility, the laboratory must evaluate the quality of the movement. The process of rating motility may vary somewhat from one laboratory to another. Normal motile sperm should have strong forward progressive motion. Automated sperm analyzers commercially available and specifically designed to evaluate semen can add specific information about motility parameters. In addition to determining percent motility, they calculate the speed at which the sperm are swimming in microns/second.

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The Disappearing Antibody: A Case Study
Transfusion Service Laboratory

The transfusion service laboratory (TS) instructed clinical staff to draw blood specimens for compatibility testing before transfusing any blood components or products.Once the blood samples were collected, the clinical staff immediately began transfusing the patient with the O Rh-negative blood.

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Unexpected anomaly

3. Do the results of the initial antibody screen support the presence of the identified antibody?No: All 3 screen cells reacted in the initial screen. Upon review, however, only Screen Cells 1 and 3 were Jk(a+); Screen Cell 2 reacted but was Jk(a-).This anomalous result was investigated by a reference laboratory. It was discovered that the patient had anti-Rd, an antibody to the low frequency antigen Radin (Rd). By chance, Screen Cell 2 was Rd-positive. Radin has a frequency of less than 0.5% in several populations tested. The screen cell manufacturer was notified. They would likely confirm that the cell was Rd-positive, make their clients aware of it, and document it in future antigrams.

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The patient's red cell eluate initially was unidentifiable, reacting weakly with only two panel cells that did not fit a pattern. Once anti-Jka was identified, a check of the eluate panel results showed that both reactive cells were Jk(a+b-) but two other JkaJka panel cells did not react.Consider the question below, then click on the answer.View Page
Literature and online resources

Literature Dutton RP, Shih D, Edelman BB, Hess J, Scalea TM. Safety of uncrossmatched type-O red cells for resuscitation from hemorrhagic shock. J Trauma. 2005 Dec;59(6):1445-9. Johnson ST, Rudmann SV,Wilson, SM. Serologic problem solving strategies:a systematic approach. Bethesda, MD: AABB, 1996.Online resourcesThe following are online examples of good practice. The information should not be used as a substitute for technical and clinical judgment. Medical and technical information becomes obsolete quickly and current sources relevant to the user's location should always be consulted. Urgent requirements for blood (Calgary Laboratory Services, Calgary,Alberta, Canada) Online resource for laboratory's clients Why is there never enough O Rh negative blood? (American Red Cross) Advice for physicians on how to help prevent shortages of O Rh negative blood Transfusion reactions: Transfusion complications (Canadian Blood Services) Education website for CBS's hospital customers REACT (Sunnybrook HSC, Toronto, ON, Canada) Pocket reference card for nurses on signs and symptoms of transfusion reactions Quick cals (online calculator of p values for Fisher's exact test) Use a one-tailed test (since we would expect an antibody to react with red cells that are positive for the corresponding antigen)

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The antibody screen is positive but the transfusion of the O Rh-negative RBCs is already in progress. What are the transfusion service (TS) laboratory's priorities in this case?Place the following procedures that will be followed by the TS in the appropriate order of priority.View Page
Crossmatch Results

These are the results of the crossmatch that was being performed in the transfusion service laboratory while the patient was receiving the two units of O Rh-negative RBCs. Cells Gel IAT* Donor I** 2+ Donor 2** 2+ Donor 3 3+ Donor 4 3+ Donor 5 2+ Donor 6 3+ * IAT = indirect antiglobulin test ** O Rh-negative RBC (Donors 3 - 6 are O Rh-positive)

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Pretransfusion Direct Antiglobulin Test Result

The laboratory obtained post-transfusion blood specimens in order to perform a serological investigation. Pretransfusion and post-transfusion DATs were performed. Patient cells DAT CC Pretransfusion 0 2+ DAT = direct antiglobulin test with polyspecific antiglobulin serumCC = IgG sensitized RBC

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Investigating weak antibodies

In this case the patient's antibody has disappeared from the plasma by adsorbing to transfused donor red cells. It is detectable but unidentifiable in the post-transfusion red cell eluate. Several trial and error procedures exist to enhance weak antibodies. Which methods will enhance the reactivity of a given antibody depend on its characteristics. Methods to investigate weak antibodies include: Use a higher plasma to red cell ratio (add more antibody-containing plasma or eluate) Increase incubation time (if consistent with manufacturer instructions, if applicable) Use enzyme-treated panel red cells (enzymes enhance IgG antibodies in Rh and Kidd blood systems but denature some antigens, e.g., Fya, Fyb, S) Try alternative antibody detection methods, e.g., if using LISS routinely, try polyethylene glycol (PEG) or column agglutination methods such as gel, providing they have been validated for use in the TS laboratory.

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The Urine Microscopic: Microscopic Analysis of Urine Sediment
Microscopic Examination

The microscopic examination was traditionally performed on all urine specimens after macroscopic exam, specific gravity and chemical tests were completed. Today, many laboratories perform a urine microscopic only if preliminary evaluation indicates the need for microscopic examination. Such laboratories must have criteria determining the specimens on which a urine microscopic will be determined. The microscopic exam is often important in detecting and evaluating renal and urinary tract disorders as well as other systemic diseases.

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An Introduction to Quantitating the Urine Microscopic

In order for a urinalysis to be useful a physician must know not only what elements are present but the quantity of each. This section will deal with counting and estimating the microscopic elements found in the urine sediment. The quantifications may vary slightly between laboratories, but each lab should have its own criteria. Quantitation may be divided into three steps: Looking for casts Counting elements Estimating elements

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Tuberculosis Awareness for Healthcare Workers
TST Interpretation and Classification

The TST interpretation depends on the measured diameter of the induration and the clinical status of the patient.An induration of 15 or more millimeters is considered positive in all persons.An induration of 10 or more millimeters is considered positive in patients in the high risk progression groups and in mycobacteriology laboratory workers.An induration of 5 or more millimeters is considered positive in the high risk infection groups.

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TB Infection Control in the Laboratory

The laboratory director is responsible for the development of a risk-based infection control plan for the laboratory.The personnel are trained in methods that minimize the production of aerosols.A respirator is used when performing procedures that can result in aerosolization outside a biological safety cabinet.Personal protective equipment specified in the infection control plan is used.Disposable gloves are worn for all laboratory procedures.

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Which of the following secondary barriers are recommended for microbiology laboratories that work with Biosafety level 3 agents (e.g., Mycobacterium tuberculosis)?View Page
Biosafety Levels

Laboratory workers who handle infectious materials in the microbiology laboratory should be aware of the work practices, safety equipment, and barriers that will protect them and others in the area from infectious agents. The Centers for Disease Control and Prevention (CDC) and the National Institutes of Health (NIH) created guidelines to assist laboratories in developing safe practices based on the infectious agents that are handled. These guidelines are referred to as Biosafety Levels 1 through 4. Each increasing number represents increased risk, requiring more stringent work practice and increasingly protective safety equipment and barriers. A copy of the Guidelines can be obtained from the CDC or accessed online at:http://www.cdc.gov/OD/ohs/biosfty/bmbl5/bmbl5toc.htm

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Biosafety Level 3

Biosafety level (BSL) 3 practices, safety equipment, and facility design and construction are applicable to microbiology laboratories that work with indigenous or exotic agents with a potential for respiratory transmission, and which may cause serious and potentially lethal infection. If the laboratory is propagating and manipulating cultures for M. tuberculosis, BSL 3 practices, containment equipment, and facilities are required. Nonaerosol-producing manipulations can be performed using BSL 2 practices, containment equipment, and facilities. At biosafety level 3, laboratory manipulations should be performed in a Class l or Class ll biosafety cabinet (BSC) or other physical containment device. Secondary barriers include controlled access to the laboratory and ventilation requirements that minimize the release of infectious aerosols from the laboratory. Secondary barriers should include self-closing double-door access and negative airflow into the laboratory. Exhausted air must not be recirculated.

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White Cell and Platelet Disorders: Peripheral Blood Clues to Nonneoplastic Conditions
Lymphocytes displayed in the photograph most likely would be called atypical or reactive. A quantitative estimate of the number of such cells may be useful using terminology such as mild (or 1+), moderate (2+) or many (3+). What percentage of the total white blood count would a report of moderate or 2+ atypical lymphocytes indicate?View Page
Assume that several other lymphocytes similar to the one in the center of the photograph are found on review of the peripheral smear. A work up for leukemia should be recommended.View Page
An electronic platelet count of 40,000/cumm was reported. Review of the peripheral blood smear(see photograph)reveals single platelets in open fields and platelet clumps. The platelet count is likely incorrect.View Page
Atypical Cells: Quantitative Estimate

Smudge cells are indicated by the arrows in this image. In some laboratories, a semi-quantitative estimate of the number of smudge cells may be made; in others, a report of "smudge cells present" may suffice. The point is that a language for reporting semi-quantitative estimates must be established for any atypical cells appearing in the peripheral blood smear. This reporting scheme must be understood by the physician in order to maximize patient care outcomes through his/her decision making process. For example, in the context of this exercise, does it make any difference to the physician if you report few or many smudge cells; or, is a report of smudge cells present sufficient? The answer to this question applies not only to smudge cells, but to the reporting of any other atypical white cells as well. An agreement must be reached between the hematology laboratory and clinical services as to how semi-quantitative estimates will impact the need for further testing in view of patient care outcomes.

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Platelet Estimate

The findings in the photograph from a peripheral blood smear would elicit a report comment of "increased platelets" of some high magnitude, such as "marked" or "4+". Estimates of platelet counts from review of a peripheral blood should be made on each smear examined. This provides a simple estimate of "high" or "low" or corroborates the value generated from an electronic cell counter. A formula for estimating platelet counts must be established in each laboratory. Following is a guideline: 5/oil power field (OPF) = 100,000/cumm; each platelet thereafter = 10,000/cumm. Thus, if an average of 10 platelets/OPF are observed, the estimated platelet count is 150,000.cumm. Such a counting scheme for platelets when clustered as in the photograph is probably not needed, as there are more than 100 platelets in the field. This translates into a platelet count of 1 million/cumm or more. This peripheral smear observation, however, would serve to corroborate an electronic platelet count of 1.2 million/ cumm.

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Additional comments on this exercise

The following pages in this presentation includes a series of white blood cell abnormalities that may be identified in a peripheral blood smear. Many of the cases will simulate the practice of a peripheral smear review by a hematology morphologist. He/she must asses what responses in patient care may be triggered by the clinician attempting to interpret the reported findings on a peripheral smearObservations of white blood cell abnormalities in the peripheral blood smear should be reported so as to direct the physician to an immediate specific diagnosis, such as: (1) atypical lymphocytes suggesting infectious mononucleosis rather than leukemia, (2) toxic granules in neutrophils as in acute infections, or atypical granules suggesting a genetic disorder, (3) an unusual mix of cells, such as too many or too few neutrophils, monocytes, or other myeloid cells, and (4) the presence of giant platelets, myelocytes, or other cells suggesting a myelodysplastic syndrome.In summary, laboratory data should be presented to clinicians in a user friendly way to promote effective decision making. The design of the data base of information must be directed toward providing clinically helpful information clearly and quickly in order to facilitate appropriate action in terms of optimizing patient care outcomes.d

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Criteria for evaluation of white blood cells and platelets

In most clinical hematology laboratories, an initial blood count is performed by an electronic instrument. Some of these instruments also produce a differential blood count, and a platelet count. Instruments that provide a 3-part differential indicate the percentage of neutrophils, lymphocytes, and a mixed field group that includes monocytes, eosinophils, basophils, immature and atypical cells. Thus, the atypical cells shown in the photograph would be counted as mixed cells and a smear review would be needed to make an identification. Instruments providing a 5-part differential count include monocytes and eosinophils. In cases where the mixed cell count is high, or there are other indications that atypical cells may be present, a hematologist's review of the smear is indicated.

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