Serological Testing In Blood Transfusion Services Salwa Hindawi Medical Director of Blood Transfusion Services KAUH 4th year medical students

4th year medical students Introduction Serology is the study of antigens and antibodies Immunohematology refers to testing done in blood banks on red blood cells All testing requires the use of Standard Precautions 4th year medical students

Antigens and Antibodies Antigens are substances recognized as foreign by the body Antibodies are proteins produced by the body in response to specific antigens 4th year medical students

4th year medical students ABO Group Four blood groups: A, B, O, or AB Almost all serum contains antibodies to ABO antigens it lacks Antibodies are crucial to safe transfusion 4th year medical students

4th year medical students Red blood cell antigens type A type AB A antigen A and B antigen B antigen H antigen (no A or B antigen) type B type O 4th year medical students

Red cell Antigens: ABO type present on RBCs, GI tract and vascular endothelium three alleles A, B, O, the A and B alleles code for glycosyltransferases specificity of the antigen is in its terminal sugar galactosamine for A galactose for B 4th year medical students

Molecular basis of ABO alleles fucose added to H-substance precursor product of A allele adds N-acetylgalactosamine product of B allele adds galactose both molecules (A and B) can be expressed on cell surface at the same time in OO individuals H substance is not modified 4th year medical students

4th year medical students Antibody in Serum Genotype Phenotype Frequency Antigen IAIA IAIO Blood Group A 41% A Anti-B IBIB IBIO Blood Group B 10% B Anti-A IOIO Blood Group O 45% Neither Anti-A Anti-B IAIB Blood Group AB 4% Both Neither Note: Caucasian frequencies given. Frequencies differ with population. 4th year medical students

4th year medical students Rh type Rh blood group antigens are present only on RBCs Five important antigens of the Rh system are D, C, E, c, and e Rh positive means that the D antigen is present (85% on the population) Rh negative means that the D antigen is absent (15% of the population) the D antigen is highly immunogenic More than 70-80% of D negative persons receiving D positive blood are expected to develop anti-D 4th year medical students

4th year medical students Rh Blood Group Alleles Multiple Alleles Weiner R1 R2 R0 Rz r rl rll ry Closely Linked Loci Fisher-Race CDe cDE cDe CDE cde Cde cdE CdE Rh+ Rh- Weiner - One complex locus, 8 different alleles Fisher-Race - Three separate loci; simple alleles, D allele most important; D = Rh+; dd = Rh- C and E loci may effect expression of Rh phenotype 4th year medical students

Other Blood Group Systems clinically significant blood group systems are Kell (K), Kidd (Jk), Duffy (Fy) and Rh (E,e,C,c) systems. antibodies are made by people who lack the antigen on their RBCs and have been exposed to RBCs containing the antigen 4th year medical students

4th year medical students Steps in Blood Banking Type and Screen (T & S): (Done for low probability of transfusion) ABO and Rh type Antibody screen Antibody identification Type and Crossmatch (T & C) (Done for high probability of transfusion) above steps plus Crossmatch 4th year medical students

4th year medical students Pre-transfusion Testing: Purpose: To select blood components that will not cause harm to the recipient and will have acceptable survival when transfused. When performed properly, pretransfusion tests will confirm ABO compatibility between the component and the recipient and will detect most clinically significant unexpected antibodies. 4th year medical students

4th year medical students Guidelines for Pre-transfusion Testing: Positive identification of recipient and recipient sample ABO group and Rh typing of recipient and donor blood. Red cell antibody detection tests for clinically significant antibodies using recipients serum or plasma. Comparison of current findings of recipients sample with record of previous patient results. Selection of components of ABO group and Rh type appropriate for the recipient. Performance of a serologic or computer crossmatch. Labeling of products with the recipients identifying information. 4th year medical students

Direct Antiglobulin Test (DAT) also called the direct Coombs test adding anti-IgG to detect IgG that is attached to the RBCs also detects C3 complement fragments on the RBC surface DAT is performed in the investigation of immune hemolytic anemia and transfusion reactions 4th year medical students

Indirect Antiglobulin Test (IAT) detects free antibodies in the serum the IAT test is performed during the antibody screen and antibody identification 4th year medical students

4th year medical students ABO GROUPING ABO grouping is the single most important serological test performed on pre-transfusion 4th year medical students

4th year medical students ABO Grouping Pt Cells Pt Serum vs vs anti -A anti-B A cells B cells A + 0 0 + 40% B 0 + + 0 11% AB + + 0 0 4% 0 0 0 + + 45% 4th year medical students

4th year medical students Type and Screen (T & S) an ABO and Rh type and an antibody screen and antibody identification are done when the patient is admitted only testing necessary if low probability of transfusion 4th year medical students

Crossmatch to Transfusion ratio (C:T ratio) blood is used more efficiently when the number of units set aside for a particular patient (crossmatched) are actually transfused. when a patient does not need blood, it is good practice to get a T& S but not a T & C C:T ratio is less than 2:1 4th year medical students

Maximum Surgical Blood Order Schedule (MSBOS) Is a guideline to order standard number of units of RBCs to be crossmatched for a specific surgical procedure, based on average use in the institution examples angioplasty T&S aortic dissection T&C 6 U 4th year medical students

4th year medical students ANTIBODY SCREENING that antibody screening should be performed in all pre-transfusion testing. As a minimum the following antigens must be expressed within the screening cell set: C, c, D, E, e, K, k, Fya, Fyb, Jka, Jkb, S, s, M, N, Lea 4th year medical students

4th year medical students Antibody Screen (IAT) recipients serum is added to 3 test RBCs (in test tubes 1 to 3 ) which have all of the important RBC antigens on them therefore if one or more of the three screening cells is positive then RBC antibody is present in the serum then do an antibody panel to identify the antibody present 4th year medical students

Antibody Identification (IAT) after the Ab screening are positive then do an antibody identification recipients’ serum is added to 10 test RBCs in a panel (test tubes 1 to 10) which contain all of the important antigens the antibody in the serum is identified 4th year medical students

4th year medical students REAGENT RED CELL PANEL NEGATIVE AUTO CONTROL SOME CELLS POSITIVE ALL CELLS POSITIVE SOME CELLS POSITIVE (SAME STRENGTH AND PHASES) (DIFFERENT STRENGTHS (DIFFERENT STRENGTHS AND/OR PHASES) SOME CELLS NEGATIVE AND/OR PHASES) AND SOME CELLS NEGATIVE Suspect Single Antibody Suspect Multiple Antibodies Suspect Multiple Antibodies -Test other selected cells to -Test selected cells to confirm and eliminate other specificities eliminate specificities -Test patient’s cells to confirm -Additional techniques may be useful they lack antigen (enzymes) -Test patient’s cells to confirm they lack antigens ALL CELLS POSITIVE WEAK REACTIVITY SEEN WITH SOME CELLS ONLY ONE CELL POSITIVE (SAME STRENGTH AND SAME PHASES) ALL SPECIFICITIES ELIMINATED (DONOR UNIT OR PANEL CELL) Suspect Antibody to High-Incidence Antigen Suspect Weakly Reactive Antibody or Suspect Antibody to Low-Incidence Antigen or (See also Multiple Antibodies) Antibody Showing Dosage Antibody to HLA Antigen -Test cells negative for high-incidence -Enhancement techniques (enzyme panel, -Test cells positive for low-incidence antigens increase amount of serum used, antigens or known strongly positive for -May need help from Reference Laboratory increase incubation time) HLA antigens for identification or confirmation -Test patient’s cells to confirm -May need to refer to Reference Laboratory -Test patient’s cells to confirm they lack antigen for identification or confirmation they lack antigen Approaches for identifying antibodies 4th year medical students

Major Crossmatch (Compatibility testing) donor RBCs (unit of blood) are tested with recipient serum to detect unexpected recipient antibodies this checks to see if the transfusion is compatible 4th year medical students

4th year medical students CROSSMATCHING Types of Cross Match: -serological IAT crossmatch. -abbreviated serological techniques immediate spin -non-serological methods electronic cross Match 4th year medical students

4th year medical students ABO Selection of Blood Components Patient ABO Type Antibody (Serum) RBCs or Granulocytes Plasma- Containing Components* Whole Blood O (45%) Anti-A andAnti-B A, B, AB, O A (40%) Anti-B A, O A, AB B (11%) Anti-A B, O B, AB AB (4%) None 4th year medical students

4th year medical students THANKS 4th year medical students