Subject: ABO Grouping
ABO GROUPING
PURPOSE
To determine the ABO type of blood donors and recipients.
PRINCIPLE
Because exposure to A and B antigens on bacteria occurs early in life, all individuals with a normal immune system who lack one or both of the antigens are expected to have the corresponding antibody after their first 6 months of life. Therefore the ABO group can be determined either by testing for the A and B antigens on the RBCs, a so-called forward or cell typing, or by testing for anti-A and anti-B in the serum, a so-called reverse or serum type. Anti-A and anti-B are typically “complete” antibodies, meaning that they agglutinate RBCs with the corresponding antigens without the need of an antiglobulin test. In vivo they are powerful hemolysins, so transfused RBCs must always be ABO compatible with the recipient’s serum. Correct determination of the ABO group of blood recipients and donors is the single most important procedure in immunohematologic testing since errors in this determination can result in fatal transfusion reactions.
For this reason it is essential to pay strict attention to sample identification, labeling of tubes, and recording, interpretation and transcription of results (see general rules, procedure #201). Whenever possible the ABO group is performed both by "forward" or cell testing (for A and B antigens) and by "reverse" or serum testing (for anti-A and anti-B); exceptions include umbilical cord blood and peripheral blood typing for infants less than 4 months of age and RBC unit confirmation, in which cases only a forward grouping is performed. When the interpretation of the forward and reverse reactions differs, the "ABO discrepancy" must be resolved by further testing. For new patients without a previous blood bank record, the ABO (and Rh) type is repeated, including preparation of a new RBC suspension, in order to detect errors.
The anti-A and anti-B reagents currently in use in this laboratory are monoclonal antibodies. These IgM reagents react strongly with most RBCs expressing weak subgroups of A and B, so ABO discrepancies caused by such blood groups will be rare. Similarly, these reagents will not detect polyagglutinable RBCs and may not detect the acquired B phenomenon.
ABO (and Rh) grouping on patients is recorded on a Patient Blood Bank Record form and filed indefinitely. Donor ABO (and Rh) grouping is recorded on the Blood Donor Processing Record and transcribed to the Blood Donor Record.
POLICIES (See also General Rules for Serologic Testing, procedure #201)
1.Both a cell (“forward”) and a serum (“reverse”) grouping will be performed for all blood donors and all patients after the neonatal period. Only forward grouping is performed for donor confirmation (repeat donor typing).
2.For new patients without a previous blood group determination the ABO group and Rh type must be repeated, including preparation of a new RBC suspension.
MATERIALS AND SPECIMENS
See General Guidelines for serologic testing, procedure # 201.
Patient Blood Bank Record, appendix 210.1
PROCEDURE
Forward (Cell) Grouping
1.Prepare a washed 35% suspension of the red cells to be tested (See procedure #202).
2.Properly label two 10x75 or 12x75 mm test tubes.
3.Place one drop of anti-A and anti-B into the appropriately labeled test tube.
4.Add one drop of the cell suspension to be tested to each of the tubes.
5.Mix and spin at the proper speed and time for the serofuge in use (noted on the side of the centrifuge.)
6.After centrifugation, examine each tube for agglutination. Record the reactions on the appropriate worksheet.
7.If the patient types as group AB and Rh type positive, prepare an Rh control tube containing one drop of 6% albuminor patient serum/plasma and one drop of the patient’s washed RBCs. Mix, spin, read, and record the results in the "6% alb" of the patient record.
Reverse (Serum) Grouping
1.Properly label two 10x75 or 12x75 mm test tubes.
2.Place 2 drops of the unknown plasma or serum (patient's or donor's) into each of the test tubes.
3.Mix reagent RBC suspension well and add l drop of the reagent A1 and B cells to the appropriate tube.
4.Mix and centrifuge at the proper speed and time for the serofuge in use.
5.Examine each tube for agglutination or hemolysis. Record the reactions and interpret the results.
Repeat Testing for New Patients
If the patient has never had a previous ABO (and Rh) groupingrepeat ABO testing must be done as follows.
1.Discard the previously used RBC suspension and prepare a new one (see procedure #202).
2.Label four new test tubes from the ORIGINAL patient specimen.
3.Repeat the testing, record the results listing “repeat” in the space for the date, and interpret as above. Compare the results and resolve any discrepancy.
INTERPRETATION OF RESULTS
The expected results for the ABO Blood Group System are:
Reaction with antibody / "Forward"Interpretation / Reaction with reagent RBCs / "Reverse"
Interpretation
anti-A / anti-B / A1 cells / B cells
0 / 0 / O / + / + / O
+ / 0 / A / 0 / + / A
0 / + / B / + / 0 / B
+ / + / AB / 0 / 0 / AB
Discrepancies Between Cell and Serum Results
If there is a discrepancy between the forward and reverse grouping, an interpretation must not be entered but the results must be recorded. Repeat both the reverse grouping and the forward grouping using a newly prepared RBC suspension. It is important at this time to record the strength of the reactions.
NOTE:Technical errors can be avoided by adherence to the "General Rules" listed in SOP #201.
1.If the discrepancy is not resolved on repeat testing, incubate the reverse type starting with 15minutes at room temperature, and re-read the tubes. If the results are interpretable, write them under the original worksheet entry indicating "Incubated" and the time.
2.If the patient is an infant less than one year old and incubation of the reverse typing has not resolved the problem, no further testing is necessary.
3.If the discrepancy is not resolved upon repetition of the test, do not enter an interpretation and proceed to procedure #223, "Investigation of an ABO discrepancy". Group O RBCs can be selected for the patient.
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