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These are the MediaLab courses that cover Wbc and links to relevant pages within the course.

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Cerebrospinal Fluid (retired 7/17/2012)
Cells Present in Normal CSF

In addition to chemical components, a few cells are also found in normal CSF. In an adult, 0 - 5 WBC/µl is considered normal. Children will have slightly higher cell counts. Up to 30 WBC/µl is within normal limits for newborns. Lymphocytes account for 60 - 100% of these cells.

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Turbidity

Spinal fluid samples are either clear or turbid. Some sources use the following rating system for turbid CSF specimens: 0 = crystal clear fluid 1+ = faintly cloudy, smoky, or hazy 2+ = turbidity clearly visible but newsprint read easily through tube 3+ = newsprint not easily read through tube 4+ = newsprint cannot be seen through the tubeTurbidity may be caused by leukocytes, erythrocytes, fungi, bacteria, amoebae, contrast media, or aspiration of epidural fat during puncture.200 leukocytes/mm3 will cause slight turbidity (1+); increased numbers of WBCs will cause increased turbidity. At least 400 erythrocytes/mm3 are needed to produce 1+ turbidity.Occasionally CSF will have an oily appearance due to the presence of substances remaining in the CSF after radiologic (x-ray) procedures have been performed.

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Normal Cell Counts

Up to 5 WBCs per microliter are present in normal adult CSF. Children have slightly higher counts, while in newborns a count of up to 30 leukocytes per microliter is within normal limits. CSF containing up to 200 WBCs or 400 RBCs per microliter may appear clear or only slightly hazy, so all specimens must be examined microscopically.

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Diluting Fluids

There are several diluents that may be used for CSF counts. Normal saline should be used to make dilutions for total cell counts. Diluting fluids for WBC counts include:crystal violet/acetic acidgentian violet/acetic acidtoluidine blue 0 and saponinThese fluids stain the white cells and lyse the red cells. The red cell count can be obtained by subtracting the white cell count from the total count. Low power (10x) may be used for the total count while the high power objective (40x) is suggested for the white cell count, especially if the white cells are to be differentiated into segs, lymphs and monocytes.

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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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True or false: if a CSF specimen is bloody due to a traumatic tap, it may be necessary to correct the CSF WBC count.View Page
Table of Normal CSF Properties

The following table lists the properties of normal CSF in adults and children: Condition Appearance Predominant Cell normal adult clear, colorless lymph 60% monocytes 30% neutrophil 2% 0-5 WBC / ul 0 RBC / ul normal neonate clear, colorless lymph 20% monocytes 70% neutrophil 4% 0-30 WBC / ul variable RBC Adapted from Saunders Manual of Clinical Laboratory Science. Craig A. Lehrmann, Ed. WB Saunders, 1998.

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Chemistry / Urinalysis Question Bank - Review Mode (no CE)
Identify the urine sediment elements shown by the arrow:View Page
Identify the urine sediment elements shown by the arrow:View Page
Which of the following cells when found upon microscopic examination of the urine would be most indicative of kidney disease:View Page

Confirmatory and Secondary Urinalysis Screening Tests
If present in the urine, which of the following may cause a false-positive urine reagent strip test for bilirubin, if the test is read visually?View Page

General Laboratory Question Bank - Review Mode (no CE)
Which of the following cells when found upon microscopic examination of the urine would be most indicative of kidney disease:View Page

Hematology / Hemostasis Question Bank - Review Mode (no CE)
Which of the two WBCs indicated by the arrows on this illustration is normally the most numerous in peripheral blood and what is its name:View Page
If greater than 50% lymphocytes were found on the peripheral blood smear of a 5 month old child you would suspect which of the following conditions:View Page
An India Ink preparation in used to identify:View Page

Normal Peripheral Blood Cells (retired 6/20/2012)
Leukocytes or White Blood Cells (WBC's)

The second group of cells are the leukocytes or white blood cells (WBC's). The leukocytes can be divided into two groups: mononuclear and granulocytic cells. Leukocytes are involved in various ways with the body's defense mechanisms. The cell shown in the upper image is a mononuclear cell, in this case a monocyte. The cell shown in the lower image is a granulocyte, in this case a neutrophil. These cells will be presented in much more detail later.

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Glossary of Terms A through M.

Antibody - A modified type of serum globulin synthesized by lymphoid tissue in response to antigenic stimulus. By virtue of specific combining sites each antibody reacts with only one antigen. Anucleate - Having no nucleus. Azurophilic granules - The well-defined large reddish granules (lysosomes) which may be present in large lymphocytes. They are called "azurophilic granules" because they stain blue with the azure stains which were originally used. Basophilic granules - Specific granules present in the cytoplasm of basophils. These granules are large and stain purple-black due to their strong affinity for basic stain. B-cell - Bone marrow derived lymphocytes which produce humoral antibodies. Biconcave - Having two concave surfaces. Cellular Immunity - The capacity of a small proportion of lymphoid population to exhibit response to a specific antigen. Chromomere - The centrally located granular portion of the platelet. Clone - A population of cells descended from a single cell. Delayed Hypersensitivity - (part of cellular immunity) that develops slowly over a period of 24-72 hours after an antigenic stimulus. It consists of an accumulation of cells around small vessels and/or nerves. Example: Tuberculin skin test reaction. Digestive Enzyme - A substance that catalyzes or accelerates the process of digestion. Eosinophilic Granules - Specific granules present in the cytoplasm of eosinophils. These granules are large, refractile spheres which stain reddish-orange due to their strong affinity for acid stain. Erythrocyte (red blood cell, RBC) - One of the elements found in peripheral blood. Normally the mature form is a non-nucleated, circular, biconcave disk adapted to transport respiratory gases. Fixed Macrophage - A phagocyte that is non-motile. Free Macrophage - An ameboid phagocyte present at the site of inflammation. Graft Rejection - A transplanted tissue that is rejected by the body's antibodies. Graft vs. Host Reaction - A complication that occurs when an implanted piece of tissue, which contains antibodies, rejects the host's tissue. Granulocyte - A leukocyte which contains granules in its cytoplasm, i.e., neutrophilic, eosinophilic, or basophilic granules. Half-life - is the length of time it takes for half of the cells circulating at a given time to leave the blood for the tissues. Hemocyte - Any blood cell or formed element of the blood. Hemostasis - A mechanism of the vascular system to arrest an escape of blood. It involves an interaction between blood vessels, platelets, and coagulation. Heparin - A mucopolysaccharide acid which, when present in sufficient amounts, functions as an anticoagulant by inhibiting thrombin. Histamine - A powerful dilator of capillaries and a stimulator of gastric secretions. Humoral Immunity - Acquired immunity produced after response to an antigenic stimulus in which B cells produce circulating antibodies. Hyalomere - the clear, blue non-granular zone surrounding the chromomere of a platelet. Immune Response - The interaction of a cell and an antigen that results in a proliferation of the cell and a capacity to produce antibodies. Isotonic Fluid - A fluid whose elements have an equal osmotic pressure. Leukocyte (white blood cell, WBC) - One of the formed elements of the blood; involved primarily with the body's defense. Lysosome - A microscopic body within cell cytoplasm; contains various enzymes, mainly hydrolytic, which are released upon injury to the cell. Megakaryocyte - A giant cell of the bone marrow from which platelets are derived. Mononuclear - A cell having a single nucleus.

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The Urine Microscopic: Microscopic Analysis of Urine Sediment
Which of the following cells may be difficult to differentiate from segmented or band neutrophils when performing a microscopic urine sediment examination?View Page
The patient is a female and the urinalysis is completed within two hours of collection.True or false? The presence of bacteria correlates with the slight turbidity and WBC result.View Page
What element is indicated by the arrow on this slide?View Page
True or false? WBCs are visible in this slide.View Page
What is the identity of the cells that are indicated by the arrow in this image?View Page
True or false? The cells that are indicated by the arrows in this urine sediment (using phase contrast microscopy) are red blood cells (RBCs).View Page
Many white blood cells (WBCs) are present in this image of urine sediment that is observed using brightfield microscopy.View Page
Renal Tubular Epithelial Cell Casts

Epithelial cell casts appear as clear cylinders containing renal epithelial cells. These casts indicate tubular damage. In the broader cast, epithelial cells appear at random: in the narrow cast, however, epithelial cells often appear in two rows. Renal epithelial cell casts may be difficult to distinguish from WBC casts especially if the casts have begun to degrade. Phase-contrast microscopy or supravital stains may aid differentiation.The epithelial cast shown in the image on the right was stained with supravital stain.

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Red and White Cells

There are many red and white cells in this high power field. There are a group of five red cells that can be identified on the lower right-hand side of the field. It is important to use the fine focus adjustment on the microscope when trying to differentiate RBCs from WBCs.Supravital stain can help to distinguish RBCs from WBCs. Two RBCs are indicated by blue arrows and a WBC is indicated by the red arrow in the lower image of a supravital-stained sediment/

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All of the following are characteristic of WBCs under high power (400X magnification) brightfield microscopy EXCEPT that they:View Page
Cell Types Observed in Urine Sediment

Cells which may be present in the urine include epithelial cells, white blood cells (WBCs) and red blood cells (RBCs). The epithelial cells in the urine may originate from any site in the genitourinary tract. It is normal to find a few epithelial cells in the sediment. WBCs may enter the urinary tract anywhere from the glomerulus to the urethra. The WBCs are mostly neutrophils. RBCs may originate in any part of the urinary tract. Normally, RBCs do not appear in the urine, although the presence of a few RBCs is not considered abnormal.

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Specimen #4 - Adult Male

The results of this specimen are abnormal but the abnormalities correlate with each other. The turbidity can be explained by the presence of bacteria and crystals. The presence of RBCs in the microscopic explains the blood found on the dipstick. The casts, bacteria and WBCs can account for the increased protein. The results may be reported.

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Counting Elements

Next the number of RBCs, WBCs, epithelial cells, parasites, and fat will be counted.Move to the center of the coverslip and examine 10 fields under high power (40X objective) brightfield. Use phase-contrast as needed. Determine the average number of each element found and record the findings as number per high power field (HPF). An abundance of any one element may be recorded as >100/HPF when 1/4 field is counted and the total field is estimated to be greater than 200/HPF.

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A 10% solution of acetic acid can be used to differentiate what from red blood cells (RBCs)?View Page
Sternheimer-Malbin Stain

The Sternheimer-Malbin (SM) stain is a commonly used supravital stain containing crystal-violet and safranin. WBC's, epithelial cells, and casts stain well with SM stain. Sedi-Strain (Clay Adams, Sparks, MD) and Kova stain (ICL Scientific) are among those commercially available. Nuclei and cytoplasm of various cells can be stained with a 0.5% solution of toluidine blue.

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Variations in White Cell Morphology -- Granulocytes
A patient has a WBC count of 4.0 x 109/L. Giant platelets, such as the one indicated by the red arrow in the image, are observed on the Wright stained peripheral smear. Blue-staining inclusions are also observed in several of the neutrophils. What is the identity of this blue-staining inclusion indicated by the black arrow in the image?View Page

White Cell and Platelet Disorders: Peripheral Blood Clues to Nonneoplastic Conditions
The cells included in the composite image were found in the peripheral blood smear of a patient with the following results:total WBC of 24.5 x 109/L. Differential count:myelocytes 1 metamyelocytes 4 band neutrophils 15 segmented neutrophils 40 monocytes 8 eosinophils 2 basophils 1 lymphocytes 29This hematologic picture is most consistent with:View Page
Criteria for Performing a Manual Review of the Smear

Automated cell counters will flag abnormal findings. The following guidelines represent typical review criteria, although specific requirements vary between laboratories. Follow the criteria established by your laboratory's procedure:Total white blood cell count <3.0 X 109/L or >12.0 X 109/LNeutrophils >80% Lymphocytes >45% or <10% Monocytes >15% Eosinophils >10% Basophils >5%A morphology review may also be indicated if the platelet count is <100 X 109/L or >650 X 109/L.

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The neutrophils illustrated in this image are representative of those seen in a female patient's peripheral blood smear. The total WBC was 28.5 X 109/L (reference interval = 4.0 - 11.0). Which of the following BEST describes the condition associated with this WBC count and the white blood cells that are present in the image?View Page
A peripheral blood smear with many myeloid cells was presented for morphology review (see image on the right). Toxic granulation and vacuoles in the neutrophil most likely represent which of the following conditions?View Page
Familial disorders: summary

Several additional familial and congenital disorders associated with atypical inclusions in WBCs are now recorded. These individual syndromes carry the following names: Fechtner, Alport, Epstein, Sebastian, and Paris-Trousseau.Fechtner syndrome( Peterson etal,Blood 65:397-406,1985)was described with 8 family members spanning 4 generations presenting with varying degrees of nephritis, deafness,and congenital cataracts. The syndrome is likely a variant of Alport syndrome with the addition of leukocyte inclusions and macrocytothemia. Several more cases involving other families have been reported. The inclusions resemble toxic Doehle bodies or those of the May-Hegglin anomaly by light microscopy, but are ultrastructurally unique.Alport syndrome is autosomal dominant, X-linked , hereditary and characterized by sensorineural deafness and hereditary nephritis. It is believed to result from abnormal glycopeptide synthesis in renal basement membranes. Recurrent hematuria and slowly progressive renal insufficiency are clinical findings. Cataracts and platelet abnormalities may be added features.Epstein syndrome is essentially Alport syndrome with the addition of macrothrombocytopenia (Seri, et al. Hum Genet 110:182-186, 2002). Neutrophil inclusions are absent in this disorder; neutrophilic inclusions are considered part of the Fechtner syndrome.The Sebastian platelet syndrome is a variant of hereditary macrothrombocytopenia combined with neutrophil inclusions that differ from Doehle bodies, but are similar to those inclusions in Fechtner syndrome. (Greinacher, et al, Blut 61:282-288, 1990).Paris-Trousseau syndrome includes large platelets containing giant alpha granules identifiable in the peripheral blood.(Breton-Gorius, Blood 85:1805,1995)

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WBC inclusions: Summary

The presence of atypical inclusions within the cytoplasm of neutrophils and other leukocytes should lead to a clinical investigation of the setting for these findings. Atypical neutrophil inclusions may be seen in the following disorders: Chediak-Higashi syndrome, May-Hegglin anomaly, Alder-Reilly anomaly, Fechtner , Sebastian, Epstein and Alport-like syndromes and in infectious and toxic conditions (in the form of Dohle bodies).Although a specific entity may not be evident from examination of the peripheral blood alone, it is important that hematology technologists include a comment reporting on the presence of these inclusions or granules. A clinical investigation with further hematologic and genetic studies may then appropriately be considered. Many of the disorders with atypical neutrophil cytoplasmic granules are also associated with platelet abnormalities, particularly giant platelets (lower image). Therefore, when atypical granules are recognized, scanning of the peripheral blood smear for atypical platelets may be revealing. These observations serve as readily identifiable markers for acquired and genetic human maladies, and as a guide for unraveling the reasons for a patient's suffering and impaired health.

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A peripheral blood smear is observed during a manual differental review. The patient is a 10 year-old boy with symptoms suggesting appendicitis and an appendectomy is being considered. The total WBC is 18.5 X 1000/uL, RBC's = 5.45 X 1M/uL, hemoglobin = 16.0 g/dL, hematocrit 48.2%.WBC differential:Segs = 53%, bands = 42% (two of which are shown in the image) monocytes = 2% lymphocytes= 2% These findings support the diagnosis of appendicitis.View Page
Case History One

A 14-year-old boy came to the physician's office with a sore throat that progressively worsened over a three-day period. His posterior pharynx was swollen, shiny and erythematous. The boy complained of pain on swallowing. His temperature was 98.5°F. A rapid direct streptococcal antigen test was positive. However, his symptoms did not subside over the next two days while on antibiotic therapy. Anorexia and nausea were persistent and compounded by a frontal headache. Cervical lymph nodes became noticeably enlarged. A complete blood count (CBC) was ordered. The results of the CBC were: WBC 11.9 x 109/L ( Reference interval= 3.8 - 9.8 x 109/L) with: 17% segmented neutrophils 5% band neutrophils 72% lymphocytes 6% monocytes All red cell findings were normal. The automated differential flagged for atypical cells, presumptively atypical lymphocytes. A peripheral blood smear was prepared. The image on the right is a representative field from the Wright-Giemsa stained smear (1000X magnification).A rapid qualitative test for infectious mononucleosis was positive. This is a case of group-A streptococcal infection superimposed on infectious mononucleosis. Symptoms subsided in three weeks following completion of the antibiotic therapy.

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