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Antigen Information and Courses from MediaLab, Inc.

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

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Laboratories Individuals

CLIA Blood Banking Review
Why would a unit of group O blood never be administered to a Bombay patient:View Page
Anti-H:View Page
Which of the following blood group antigens are most susceptible to destruction by the action of enzymes:View Page
Which of the following group B antigens is generally associated with a mixed field reaction:View Page
The term used to describe patients with absence of Rh antigens is:View Page
The classification of Du refers to:View Page
All of the following cellular antigens are important to an immunohematologist except:View Page
Deglycerolized red cells are most effectively used to:View Page
In HDN which of the following antigen-antibody reactions is occurring:View Page
Which of the following antibodies is detected primarily in the antiglobulin phase of the crossmatch:View Page
Which of the following blood group antigen-antibody reactions is enhanced by using enzymes:View Page
Which of the following blood groups reacts least strongly with Anti-H:View Page
If an Rh negative patient is administered a unit of R1R1 packed red cells, which one of the following antibodies would be most likely to develop:View Page
Which of the following best describes the direct antiglobulin test principle:View Page
An Rh positive individual is always positive for which of the following antigens:View Page
Avidity is best described by which of the following statements:View Page
Which of the following options gives in order from most to least important, the factors you would use to select blood for a transfusion:View Page
Which of the following statements is not true about the Lewis blood group:View Page
Which of the following is not a major Rh antigen:View Page
Which of the following best describes the primary function of antibodies:View Page
The most definite indication that a patient has been sensitized to a specific red cell antigen is:View Page
DR antigens are found in which of the following systems:View Page
Patients with antibody to the following antigen are immune to Hepatitis B:View Page
Which of the following statements best describes Rh antibodies:View Page
Which of the following Rh antigens is found the highest frequency in the Caucasian population:View Page
Which of the following set of conditions would preclude hemolytic disease of the newborn as a result of ABO incompatibility:View Page
Pre-transfusion testing should include all of the following except:View Page
To detect the presence of blocking antibodies fixed on the red cells of a newborn infant:View Page
Which of the following Rh antigens is found the highest frequency in the Caucasian population:View Page
Unexpected positive reactions encountered during forward ABO typing may be due to:View Page
The antigen marker most closely associated with transmission of HBV infections is:View Page
HLA-A and HLA-B antigens can be detected using which of the following techniques?View Page
Which of the following antigen groups is closely related to the ABO system:View Page
Proteolytic enzyme techniques may be useful in identifying which of the following antigen groups:View Page
Which of the following red blood cells contain the most H antigen:View Page
A patient's serum reacts with all reagent red cell samples. The autocontrol is negative. An alloantibody to a high incidence antigen is suspected. Which of the following would be most likely to be a compatible donor:View Page
The two or three reagent cells used for antibody screening will detect which of the following:View Page
HLA antigen testing may be used for all except the following:View Page
ABO blood groups were discovered by:View Page

CLIA General Laboratory Review
An increase in CEA levels is most closely associated with which of the following organs:View Page
The reaction that occurs when a soluble antigen is mixed with its specific antibody is termed:View Page
Which of the following immunoglobulin classes is chiefly responsible for the degranulation of mast cells and basophils:View Page
A decrease in which of the following in AIDS patients results in increased susceptibility to infection:View Page
Match terms to IgG molecule illustration:View Page

CLIA Hematology / Hemostasis Review
An India Ink preparation in used to identify:View Page

CLIA Microbiology / Serology Review
VDRL is an example of which of the following types of tests:View Page
Which of the following substances produced by Group A Streptococci is responsible for producing type specific immunity:View Page
Which of the following tests would be used to directly document the presence of a specific organism in a clinical specimen:View Page
Which of the following hepatitis antigens is most directly linked to transmission of HBV:View Page
A decrease in which of the following in an AIDS patient are associated with increased susceptibility to infection:View Page

Current Topics in Clinical Microbiology
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
Beta hemolytic colonies grew from the blood culture bottle after 18 hours incubation (see photograph). The following tests would be helpful in making a preliminary identification:View Page

Fundamentals of Molecular Diagnostics
Overview

To aid in the diagnosis of disease or identification of infectious agents, clinical laboratorians use a variety of methodologies to assist them. Knowing what to look for, or the right question to ask, is vital to obtaining the correct answer. Many diseases and agents have unique causes. The cause of the condition then becomes the "target" to be identified and perhaps even quantified. For example: If Patient A is suspected of having disease X, and disease X requires treatment, it is necessary to prove that disease X exists within patient A. We must know something about what causes disease X; is disease X an antigen, a bacteria, a viral particle, a missequenced piece of DNA?Once the target of interest (in this case Disease X) has been identified, the clinical laboratorian can choose the methodology most appropriate to answering the question, "Does disease X exist within Patient A?"

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HIV Safety for Florida
Before a HIV antigen or antibody test can be ordered, informed consent must be obtained.View Page
Informed Consent

Informed consent must be obtained first before a HIV antigen or antibody test can be ordered.Informed consent must be preceded by: Explanation of the test subject's right of confidentiality. Notification that a positive HIV test result will be reported to county health department with enough information to possibly identify the test subject. Availability and location of sites where anonymous testing is performedInformed consent can be given by a legal guardian or other person authorized by law when the test subject is: not competent incapacitated a minor

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Introduction to Quality Control
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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Introduction to the ABO Blood Group System
Match the blood type on the left with the appropriate description on the right.View Page
An individual with type AB blood will demonstrate the complete absence of which of the following antigen sites?View Page
Reverse typing is done using known antisera to detect ABO antigens present on the patient's red blood cells.View Page
The History of the ABO System

In 1900, a German scientist, Karl Landsteiner, discovered that blood groups differ from one individual to another. He took blood samples from five associates and himself, allowed them to clot, and then separated the serum from the cells. Landsteiner found that when he mixed the serum and red cells from different individuals, some samples clumped and some didn’t. Our present day classification of the ABO system is based on Landsteiner’s realization that agglutination occurred because of highly reactive antigens present on the red blood cell which corresponded to antibodies present in the serum. Landsteiner isolated and named the red cell antigens “A” and “B” and the corresponding antibodies “Anti-A” and “Anti-B.” If the red cells contained neither antigen, he called these cells “O”, representing zero antigens present. The fourth type of red cells, “AB”, was discovered in 1902 by Von Decastello and Sturli, associates of Landsteiner. “AB” cells contained both A and B antigens on their surface.

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The History of the ABO System (cont.)

Landsteiner, knowing that none of his subjects had been immunized, realized that “natural” antibodies must develop which are directed against antigens not present on the red cells. Individuals with “A” antigens on their red cells had sera containing “Anti-B” antibody. Individuals with “B” antigens had sera containing “Anti-A.” “AB” individuals had sera with no ABO antibodies present and “O” individuals’ sera contained “Anti-A” and “Anti-B.” Sera from group O individuals may contain a separate antibody, “Anti-A,B.” Anti-A,B possesses serologic activity not found in mixtures of Anti-A and Anti-B. Anti-A,B sera will agglutinate A, B, and AB cells. It is particularly useful in detecting weak A and B antigens. See the table on the next page.

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Table 1: ABO Blood Group System

Antigen on Red Cells Antibodies in Serum ABO Blood Group A Anti-B A B Anti-A B Neither A nor B Anti-A, Anti-B, Anti-A,B O A and B Neither Anti-A nor Anti-B AB

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Table 2: Testing the Patient Red Cells with Known Antisera (Forward Typing)

In routine practice, specially prepared blood grouping sera containing anti-A, anti-B, (and optionally anti-A,B) are used to identify the four types of red cells. These sera will agglutinate cells with the corresponding antigen. This is called forward typing. ABO Blood Group Patient Red Cells Tested with Known Antisera Anti-A Anti-B Anti-A,B A 4+ 0 4+ B 0 4+ 4+ O 0 0 0 AB 4+ 4+ 4+ + = agglutination (graded 1+ to 4+)0 = no agglutination

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Table 3: Testing the Serum with Known Red Cells (Reverse Typing)

It has been demonstrated that antibodies occur predictably in the sera of all normal adults in association with the ABO antigens. Demonstration of these antibodies is therefore necessary for definitive classification of an individual’s ABO cell type. The individual’s serum is therefore tested against reagent red cells containing known antigens. Patient ABO Blood Group Patient Serum Tested with Known Reagent Cells A Cells B Cells A 0 4+ B 4+ 0 O 4+ 4+ AB 0 0 + = agglutination (graded 1+ to 4+)0 = no agglutination or hemolysis

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Importance of Understanding the ABO System

While the predictability of ABO antibodies in persons lacking the corresponding antigen makes the ABO blood group system an easy one for testing purposes, it can be treacherous as far as transfusion is concerned. If a patient receives cells containing A or B antigens and his/her serum contains the corresponding antibody, the donor cells will be destroyed almost immediately with severe and sometimes fatal transfusion reaction. It is, therefore, of utmost importance to thoroughly understand the ABO blood group system. Compatibility of the ABO system is essential for all other pre-transfusion testing.

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Epitopes

It is also important to note that in addition to red cells, ABO antigenic determinants (epitopes) are found in many tissues, body fluids, and other cells including endothelial cells and platelets. Because ABO antigens are so widely expressed, ABO antigens are also a major consideration in solid organ and bone marrow transplants.

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Why does agglutination (clumping) sometimes occur when red cells from one individual are mixed with serum from another?View Page
Match the blood types in the drop down boxes with the characteristics on the right.View Page
Galactose and ABO Antigen Precursor Substance

Specific sugars, attached to the red cell membrane in unvarying linkage conformations, determine ABO antigenic activity. Galactose resides at the end of this specific sugar chain. This configuration constitutes the ABO antigen precursor substance.

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Fucose

Another sugar, fucose, must be attached to the galactose in a specific configuration for further antigen development to take place. This “galactose-plus-fucose” configuration has antigenic activity called “H”.

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"A" Antigenic Activity

Without H substance (also known as H antigen or substance H), there is no way for additional sugar attachment to take place. Additional sugar attachment is necessary for the development of A and B antigenic activity. Therefore, without substance H there can be no A and B antigens developed. Once substance H is developed, the addition of the sugar N-acetylgalactosamine to the terminal position of the chain gives the molecule “A” antigenic activity.

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The H gene

Three separate loci (ABO, Hh, and Se) contain the genes that control the location and occurrence of the A and B antigens. Hh and Se genes are closely linked on chromosome 19. The precursor substance is acted upon by the H gene and is converted to H substance. The product of the H gene is an enzyme fucosyltransferase, responsible for attaching fucose to the terminal galactose of the precursor substance on the RBC membrane and thus forming H substance. There are only two recognized alleles at this locus: the active form, H, and an amorph, h. The H gene is a high-incidence gene. People who inherit hh are extremely rare. Since the h gene is amorphic, it does not act on the precursor substance.

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A, B, and O Genes

The ABO locus is on chromosome number 9. There are three major allelic genes and numerous rare genes. The three principle genes are A, B, and O. The A gene determines the product N-acetylgalactosaminyltranferase activity. The B gene determines galactosyltransferase activity. The O gene does not produce a functional enzyme. The enzyme products of the A and/or B genes act on H substance to convert it to A and/or B antigens. Not all H substance is converted; thus, all cells normally contain some H substance along with the A and/or B antigens. If both the A and B genes are present, some H antigen sites are converted to A antigen and other H antigen sites are converted to B antigen. (A single antigen site does not have both A and B antigens.) The O gene is an amorph and doesn’t act on H substance, therefore group O cells contain only H substance. See the diagram on the next page.

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Bombay Blood Group Genes

As mentioned previously, the A and B genes cannot act directly on the precursor substance. Thus, since individuals with the Bombay phenotype have only the precursor substance and no H antigen, they cannot have A or B antigens, even if they have the A and/or B gene.

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Inherited Genes

The A, B, and H antigens, like many other blood group antigens, are the expression of genes inherited from the previous generation. If the antigen is demonstrated, the gene controlling it must have been inherited from one or both of the parents.  As previously mentioned, the genes A, B, and O are allelic genes. Assuming the production of H substance, these three genes, in various possible combinations of two, account for the four recognized ABO groups: A, B, AB, and O. Each individual inherits two ABO genes, one from each parent, and these genes determine which ABO antigen will be present on that individual’s red cells. These genes exhibit co-dominance, meaning that if both A and B genes are present, both will be expressed.

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Deducing the Gene

The presence of A and/or B antigen on the red cells can be recognized by serological tests with the appropriate antisera so that the presence of the gene that controls its production can be deduced in the absence of both A and B genes (when no A or B antigen is present on the red cells).

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Genotyping Through Genetics

Those who type as group O must have two O genes present (since both the A and B genes would have produces recognizable antigens, neither of which is present on group O cells). Therefore, in the case of an AB individual or an O individual, we can tell exactly which genes are present, or a genotype. Typing that show persons to be group A or group B reveal only one gene product and thus only a phenotype can be determined. Persons of phenotype A can be genotype AA or AO , while those of phenotype B can be genotypically BB or BO. Family studies may be done to determine the genotype of an A or B individual. Fore example, if the mating of one A and one O parent produced a group O child, the second gene present in the A parent must have been O since the child has inherited one O gene from each parent.

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How many gene loci regulate red cell ABO antigen development?View Page
Which of the following is true of Bombay cells?View Page
ABO Antibodies

In most other blood group systems, antibody may be formed after an individual has been immunized by an antigen that is missing from his or her red cells; perhaps as the result of pregnancy or transfusion. In the ABO system, when the antigen is missing from the cells, the corresponding antibody will predictably be found in the serum and must be found before determining the ABO type. There are few exceptions to this rule and any exception must be explained before the true ABO blood type can be determined.

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Anti-A and Anti-B Development

It is possible that since anti-A and anti-B develop so predictably, without a recognizable immunizing event, that they are “naturally” occurring. Their production is thought to be stimulated by bacteria which have been shown to contain substances that are chemically similar to human A and B antigens. (Anti-A and anti-B are absent in germ-free animals.) Except for the rare hh individuals who lack H substance, everyone has some H in their cellular makeup.

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Immunoglobulin

The predominant immunoglobulin class for the B antibodies produced by individuals with group A phenotype and the A antibodies produced by individuals with group B phenotype is IgM. Small quantities of IgG may also be present. IgG is the predominant immunoglobulin for the anti-A and anti-B antibodies found in individuals with group O phenotype. Infants of group O mothers are at higher risk for hemolytic disease of the newborn (HDN) than those born to mothers with group A or B because IgG immunoglobulins readily cross the placenta. IgM molecules do not readily cross the placenta because of their larger size. It is important to note that immune antibodies are usually IgG. Both naturally occurring and immune ABO antibodies are critically important in transfusion since both sensitize and usually hemolyze red cells with the corresponding antigen.

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Anti-A and anti-B are stimulated by bacteria which have been shown to contain substances that are chemically similar to human A and B antigens.View Page
Strength of the A Antigen

The strength of the A antigen can vary considerably, and although most A cells react strongly with anti-A and anti-A1B, some cells have been found that are very weakly reactive. The blood group has been divided into subgroups and is classified not only by the strength of the A antigen but also by certain other serologic characteristics.

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A1 and A2

The most common classifications are A1 and A2. These account for over 99% of group A bloods. Of this 99%, A1 compromises approximately 80%. Commercial anti-A typing serum does not differentiate between A1 and A2 cells. A1 cells contain “A” antigen and “A1” antigen. A2 is not really a unique antigen. It is thought to be simply “A” antigen with no “A1” antigen. Several preparations are available that will react with A1 cells, but not other subgroups of A. An extract of the seeds of the plant, Dolichos biflorus has specific anti-A1 activity. “Absorbed anti-A” serum can also be prepared. To do this, the anti-A from group B people is absorbed with A2 cells. Anti-A is removed and a second antibody that reacts only with A1 cells remains. Anti-A1 can also be found as a separate antibody in the sera of A2 and A2B individuals.

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Inherited Antigens

A subgroup antigens are inherited, as are other ABO antigens with A1 being dominant over A2. A phenotypically A1 individual may be genotypically A1O, A1A1, or A1A2. A phenotypically A2 individual may be genotypically A2A3. These alleles are passed to offspring in the same manner as other ABO antigens. Weak variant forms of the B antigen (B3, Bx, and Bel) exist but are so rare that they do not warrant discussion here.

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Why Knowledge of A Subgroups Is Important For Laboratorians

For the most part, subgroups are merely of academic interest, but occasionally they present clinical problems. The antigen may be so weak that it is not detected and the red cells are mistyped as group O. This is especially dangerous if the cells are those of a donor. Problems may arise because the serum of an A2 or A2B, A3 or Ax individual might contain anti-A1. This antibody may be detected in serum typing and cause confusion. You would not expect to find a person with A antigen on his red cells and anti-A in his serum. Anti-A1 is produced by about 1-2% of group A2 persons and about 25% of group A2B persons. Subgroups may be determined by reactions with antisera as seen in the table on the next page.

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Why may the presence of A subgroups cause ABO typing discrepancies?View Page
Forward Typing

Forward typing is done using known antisera to detect ABO antigens present on the patient’s red cells. In the tube test, known antisera and patient cells are placed in labeled test tubes, centrifuged, and observed for agglutination. Each manufacturer has specific instructions for its own antisera, detailing the percent of cell suspension, number of drops of cell suspension versus number of drops of antisera, and the rate and length of centrifugation. Though the details differ, the theory behind the tests is the same.

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Testing the Red Cells With Known Antisera

Patient Red Cells Tested With Known Antisera ABO Antigens Present on Red Cell Anti-A Anti-B Anti-A,B 4+ 0 4+ A 0 4+ 4+ B 0 0 0 Neither A nor B 4+ 4+ 4+ A and B + = agglutination (graded 1+ to 4+) 0 = no agglutination or hemolysis

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Which of the following statements best describe forward typing?View Page
Which of the following best describes reverse typing?View Page

Laws and Rules of the Florida Board of Clinical Laboratory Personnel
Description of Specialties (1)

Specialists in microbiology perform testing to diagnose and stop the spread of infectious organisms, including bacteria, viruses, and parasites. Specialists should be able to isolate and identify a wide variety of these organisms. Testing procedures include direction examination and antigen detection methods. Specialists in serology and immunology measure antibodies to infectious organisms. Specialists should be familiar with all serology techniques (except those specific to immunohematology). This specialty includes all lab procedures performed in the specialty of histocompatibility. Specialists in hematology must be able to identify and evaluate cells in blood and bone marrow and identify disorders of these cell. Specialists should be familiar with routine and special tests to determine the number, morphology, and function of cells in body fluid.

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Mycology: Hyaline and Dematiaceous Fungi
Several saprophytic, hyaline molds have microscopic characteristics that mimic the mold forms of the dimorphic fungi (Histoplasma capsulatum, Blastomyces dermatitidis, etc.). Each of the following can be used to differentiate the saprophytic from the dimorphic fungi except:View Page

Mycology: Yeasts and Dimorphic Pathogens
Of the following responses, the one observation that would rule out cryptococcosis as the cause of meningoencephalitis is:View Page
This photomicrograph is a representative field of a Wright-Giemsa-stained bone marrow aspirate in which a pair of budding yeast cells is seen centrally (arrows). Based on the appearance of these yeast cells, what other test would you expect to be positive?View Page

Normal Peripheral Blood Cells
Humoral Immunity

Humoral immunity involves the production of antibodies (immunoglobulins), and is brought about by lymphocytes which we call B-cells. B-cells are bone-marrow derived lymphocytes. After B-cells are stimulated by an antigen, they proliferate and transform into plasma cells which produce specific antibodies.

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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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Phlebotomy
Acute hepatitis panel

Acute hepatitis panel: Hepatitis A antibody (IgM) Hepatitis B core antibody, IgM (HBcAb) Hepatitis B surface antigen (HBsAg) Hepatitis C antibody

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Obstetric panel

CBC Hepatitis B surface antigen Antibody, rubellaSyphilis test (RPR) Antibody screen Blood type, Rh and ABO

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White Cell and Platelet Disorders: Peripheral Blood Clues to Nonneoplastic Conditions
Case history

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 ,shiney and erythematous. The boy complained of pain on swallowing. His temperature was 98.5F. 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. The results of the CBC were: WBC 11.9/mm3 with 17% segmented neutrophils, 5% bands, 72%(60% atypical--see photograph)lymphocytes and 6%monocytes. All red cell findings were normal. A monospot test was positive. This is a case of group-A streptococcal infection superimposed on infectious mononucleosis. Symptoms subsided in 3 weeks following completion of the antibiotic therapy.

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