BB-3-ABO BLOOD GROUP.pdf

IMMUNOHEMATOLOGY ABO (001) BLOOD GROUP ABO BLOOD GROUP Most important of all blood groups in transfusion practice The three genes that code for A, B, O are located at the long arm of chromosome 9 Histoblood group ABO ALLELE THEORIES 1. Three Allele Theory (Bernstein) PHENOTYPE POSSIBLE GENOTY...

IMMUNOHEMATOLOGY ABO (001) BLOOD GROUP ABO BLOOD GROUP Most important of all blood groups in transfusion practice The three genes that code for A, B, O are located at the long arm of chromosome 9 Histoblood group ABO ALLELE THEORIES 1. Three Allele Theory (Bernstein) PHENOTYPE POSSIBLE GENOTYPES A AA, AO B BB, BO AB AB O OO ABO ALLELE THEORIES 2. Four Allele Theory (Thomson) PHENOTYPE POSSIBLE GENOTYPES A1 A1A1, A1O, A1A2 A2 A2A2, A2O A1B A1B A2B A2B B BB, BO O OO Frequencies (%) of ABO Phenotypes Phenotype Whites Blacks Native Americans A 42 27 16 28 B 10 20 4 26 AB 4 4 <1 5 O 44 49 79 41 Asians LANDSTEINER'S LAW 1. The antigen on the RBC determines the blood group 2. The corresponding antibody is never found in the individual's serum 3. The opposite antibody is always present in the individual's serum RBC Blood Cell Type A B AB Antigen in RBC Antibody in Plasma O NONE NONE ABO ANTIGENS Found on RBCs, lymphocytes, platelets, tissue cells, bone marrow and other organs be secreted by tissue cells if the appropriate genes are present Developed in utero at 5-6 weeks of gestation; full expression occurs between 2-4 years of age TYPE 1 TYPE 2 Linkage BETA-1,3 BETA-1,4 Origin Plasma Seen on erythrocytic precursors Controlling genes H, A, B, Se, and Lewis H, A, and B Precursor Oligosaccharide Chains PROTEIN CERAMIDE GLUCOSE GALACTOSE N-ACETYLGLUCOSAMINE GALACTOSE A OR B ANTIGEN H ANTIGEN PRECURSOR SUBSTANCE RBC PROTEIN GENE ---> ENZYME ---> ANTIGEN FORMATION OF H-ANTIGEN SUBTYPES OF H-ANTIGEN: a. H1 b. H2 c. H3 d. H4 CERAMIDE GLUCOSE GALACTOSE N-ACETYLGLUCOSAMINE L-FUCOSE GALACTOSE Gene Glucosyltransferase Sugar Acceptor Ag H a-2-L-fucosyltransferase L-fucose Precursor H-Ag H Ag A H Ag B H Ag AB A B AB O a-3-N-acetylgalactosaminyltransterase N-acetyl-D-galactosamine a-3-D-galactosyltransferase D-galactose a-3-N-acetylgalactosaminyltransterase N-acetyl-D-galactosamine a-3-D-galactosyltransferase D-galactose O O > A2 > B > A2B > A1 >A1B Comparison of ABH Antigens on RBCs and in Secretions ABH Antigens on Red Cells A, B and H Soluble Substances RBC antigens can be glycolipids, glycoproteins, or glycosphingolipids Secreted substances are glycoproteins RBC antigens are only synthesized on Type 2 Secreted substances are primarily precursor chains synthesized on Type 1 precursor chain Enzyme: a-2-L-fucosyltransferase by Se gene Se enzyme Enzyme: a-2-L-fucosyltransferase by H gene Secretor: SeSe, Sese (78%) Nonsecretor: sese (22%) ABO ANTIBODIES BLOOD GROUP ANTIBODY PRODUCED A ANTI-B B ANTI-A AB NONE O ANTI-A, ANTI-B, ANTI-A,B CHARACTERISTICS IgM Naturally occurring antibodies React at room temperature (20-24°C) Cannot cross the placenta Mostly IgM (some lgG) Anti-A,B Predominantly IgG (with small portions as IgM) Immune antibodies React at 37°C Can cross the placenta ABO TYPING BLOOD GROUP ANTIGEN A A B B AB AB O NONE ANTI-A,B REACTION WITH ANTI-A ANTI-B Checks for the reaction of anti-A and anti-B Detects weak subgroups of A and B ANTI-A,B INTERPRETATION DETERMINATION OF SECRETOR STATUS PRINCIPLE: HEMAGGLUTINATION INHIBITION DETERMINATION OF SECRETOR STATUS PRINCIPLE: HEMAGGLUTINATION INHIBITION FORWARD GROUPING Characteristics of Routine Reagents useful for ABO Testing. Anti-A Reagent Monoclonal antibody Highly specific IgM Clear blue colored reagent Blue dye: Bromphenol blue, Thymol blue, Patent blue Anti-B Reagent Monoclonal antibody Highly specific IgM Clear yellow colored reagent Yellow dye: Acriflavin, Tartrazine yellow PRESERVATIVE: Sodium azide (0.1%) "AB-BY" LECTINS For ABO antigens For antigens causing polyagglutination Arachis hypogaea: peanut extract Glycine soja: soy bean extract LECTINS SEROLOGIC SPECIFICITY FOR ABO ANTIGENS Ulex europaeus Anti-H Lotus tetragonolobus Anti-H Dolichos biflorus Anti-A, Anti-Tn, Anti-Cad Helix pomatia Anti-A, Anti-Tn, Anti-Cad Griffonia simplicifolia (Bandeiraea simplicifolia) Anti-B, Anti-T FOR ANTIGENS CAUSING POLYAGGLUTINATION Arachis hypogaea Glycine soja (Glycine max) Salvia sclarea Salvia horminum Anti-T, Anti-Tk Anti-T, Anti-Tn, Anti-Cad Anti-Tn Anti-Tn, Anti-Cad (separable) FOR OTHER BLOOD GROUP SYSTEM Iberis amara Anti-M Vicia graminea and Bauhinia species Anti-N REVERSE TYPING Reverse typing --› Antibody Typing Serum/Plasma (sample) + RCS (reagent) BLOOD GROUP ANTIBODY REACTION WITH A1 CELLS A B AB O B CELLS INTERPRETATION 4+ One solid aggregate, clear background 3+ Several large aggregates, clear background 2+ Medium-sized agglutinates, clear background 1+ Small agglutinates, turbid background W+ Tiny agglutinates, turbid background 0 MIXED FIELD No agglutination or hemolysis A type of agglutination pattern in which numerous small clumps of cells exist amid a sea of free cells BOMBAY PHENOTYPE (H NULL) hh genotype No H antigens formed; therefore, no A nor B antigens formed Phenotypes as blood group O Anti-A, anti-B, anti-A,B and anti-H present in the serum Can only be transfused with blood from another Bombay (Oh) FORWARD TYPING REVERSE TYPING PHENOTYPE anti-A O Oh anti-B anti-H A cells B cells O cells ABO SUBGROUPS Reaction with PHENOTYPE Ag Present Population Freq A1 A A1 80% (+) (+) A2 A 20% (+) (-) Note: 1-8% of A2 25% of A2B Anti-A Anti-A1 (anti-A + anti-A1) Dolichos biflorus -----> can form Anti-A1 FOWARD TYPING REVERSE TYPING CONTROLS ANTI-A ANTI-B ANTI-A,B A1 CELLS B CELLS AUTO O CELLS 3+ 0 4+ 0 3+ 0 0 W+ 0 2+ 0 3+ 0 0 W+ 0 2+ 1+ 3+ 0 0 ANTI-A1 ANTI-B OTHER SUBGROUPS OF A Phenotypes Description A3 Mixed field agglutination with anti-A and/or anti-A,B Ax Weak agglutination with anti-A,B only Aend <10% red cells show very weak mf agglutination Am No agglutination with anti-A and anti-A,B Secretors demonstrate quantities of A substance in saliva Ay No agglutination with anti-A and anti-A,B Secretors contain small amount of A substance in saliva Ael No agglutination with anti-A and anti-A,B Secretors contain only H substance and no A substance in saliva SUBGROUPS OF B Phenotypes Description B3 Mixed field agglutination with anti-B and/or anti-A,B Bx Agglutination with anti-A,B (wk/0 with anti-B) Bm No agglutination with anti-B and anti-A,B Secretors demonstrate quantities of B substance in saliva Bel No agglutination with anti-B and anti-A,B Secretors contain only H substance and no B substance in saliva ABO TYPING DISCREPANCIES: TECHNICAL ERRORS: Incorrect or inadequate identification of blood specimens, test tubes, or slides Cell suspension either too heavy or too light Clerical errors or incorrect recording of results A mix-up in samples Missed observation of hemolysis Failure to add reagents Failure to add sample Failure to follow manufacturer's instructions Uncalibrated centrifuge Overcentrifugation or undercentrifugation Contaminated reagents Warming during centrifugation ABO TYPING DISCREPANCIES: GROUP 1 DISCREPANCIES: WEAKLY REACTING OR MISSING ANTIBODIES Newborns Elderly patients Patients with leukemia demonstrating hypogammaglobulinemia (e.g., CLL) Patients with lymphomas demonstrating hypogammaglobulinemia (e.g., malignant lymphomas) Patients using immunosuppressive drugs that yield hypogammaglobulinemia Patients with congenital agammaglobulinemia Patients with immunodeficiency diseases Patients with bone marrow transplantations (develop hypogammaglobulinemia) Patients whose existing ABO antibodies may have been diluted by plasma transfusion or exchange ABO subgroups GROUP 2 DISCREPANCIES: WEAKLY REACTING OR MISSING ANTIGENS Subgroups of A and/or B Leukemia (e.g., AML) Hodgkin' disease Excess amounts of blood group-specific soluble substances (BGSS) present in the plasma in association certain diseases Acquired B phenomenon: Occurs when bacterial enzymes modify immunodominant blood sugar A sugar (N-acetyl-D-galactosamine) into D-galactosamine (similar to Dgalactose) which cross reacts with anti-B antisera Resolution: Use monoclonal anti-B clone (ES4) Treat RBCs using acetic anhydride Antibodies to low incidence antigens B(A) phenomenon CERAMIDE GLUCOSE GALACTOSE N-ACETYL-D-GLUCOSAMINE L-FUCOSE GALACTOSE QUESTION: Which ABO typing result presents a patient with an *Acquired B" phenomenon? Anti-A1 Anti-B A cells B cells a + 0 0 + b + + 0 0 c + + + 0 d + + 0 + e + + + + GROUP 3 DISCREPANCIES: PROTEIN OR PLASMA ABNORMALITIES RESULTING TO ROLEAUX FORMATION Elevated levels of globin from certain disease states (E g. muple myeloma, Waldenstrom's macroglobulinemia) Elevated levels of fibrinogen Plasma expanders Wharton's jelly GROUP IV DISCREPANCIES: MISCELLANEOUS POLYAGGLUTINATION POLYAGGLUTINATION Refers to the agglutination of altered RBCs by large proportion of ABOcompatible adult human sera Tn: marker for pre-leukemic state P. mirabilis infection ---> Acquired A Cad: resistance to P. falciparum Confirmation: RBCs Agglutination with MOST adult sera NO agglutination with cord sera PANAGGLUTINATION Patient's RBCs are agglutinated by ALL including OWN serum Lectins for Polyagglutination: Tn Cad T Tk Arachis hypogaea - - + + Salvia sclarea + - - - Salvia horminum + + - - Glycine soja + + + - Cis-AB Phenotype Cis-AB refers to the inheritance of both AB genes from one parent carried on one chromosome and an O gene inherited from the other parent COLD ANTIBODIES: RBC (sample): Wash with saline at 37°C ---› retype 0.01 M Dithiothreitol (DTT) ---> disperse IgM Serum (sample): Warmed at 37°C ---> read results at 37°C (Pre-Warmed Technique) IMMUNOHEMATOLOGY END Thank you for listening