In Focus
What is the added value of the reverse group in ABO testing?
Omit or not omit?
ABO blood group determination is unique among immunohaematology tests because it confirms the absence of specific antigens by detecting the presence of naturally occurring antibodies against the absent A and B antigens.
This method of so-called “reverse grouping” has been standard practice for over a century and is obligatory for ABO blood group determination. The necessity of reverse grouping for ABO typing was discussed during a recent Working Party meeting break-out session. The possibility of omitting reverse grouping, hence only performing ABO antigen typing, was considered.
Importance of Reverse Grouping in Different Contexts
It is essential to distinguish between ABO blood group determination in patients and donors due to the significantly different impacts of false positive or negative ABO typing results for donor versus patient categories. Omitting reverse grouping can result in potential discrepancies between antigen and antibody identification going undetected. Some of these situations can potentially affect the reliability of the results.
Potential Discrepancies in Antibody Reactions in Patient Testing
Absent or weak reactions in ABO antibody testing are not uncommon. This can occur in young children, the elderly, or following bone marrow transplants (BMT) or stem cell transplants (SCT). It can also be observed, though rarely, in cases of ABO blood group chimerism. Omitting testing for the presence of anti-A and anti-B in such cases would not affect the ABO grouping result. In this scenario reverse grouping is not of added value.
Omitting reverse grouping can save a substantial amount of unnecessary work. Many discrepancies arise from absent reactions, which, after extensive additional testing, result in the same outcome as antigen determination alone. Unexpected positive or extra reactions in reverse grouping, such as A1 test erythrocytes showing a positive test result whereas the patient is A or AB antigen positive, or B test erythrocytes showing a positive test result whereas the patient is B or AB antigen positive, are also notable. These reactions are typically caused by IgM class antibodies, reactive at room temperature, which could be alloantibodies (e.g., anti-M, anti-N, anti-Lea, anti-Leb, anti-P1) or autoantibodies (e.g., anti-I, anti-H, anti-HI, or pan-reactive). These antibodies are in general not problematic for transfusions if they are not detected in the routine red blood cell antibody screen.
Antibodies with anti-A1 specificity could pose problems since they are not detected in a standard antibody screen. These antibodies are usually reactive at lower temperatures but can sometimes be active at 37°C. Although these antibodies rarely cause transfusion reactions, there have been isolated reports of such occurrences.
When performing a plasma transfusion, information about the recipient's anti-A and anti-B antibodies may be of importance rather than ABO antigen typing results. For instance, if a subgroup of A is not detected, without reverse grouping, a patient may be classified as blood group O. If this patient is transfused with group O plasma, which contains anti-A, this could bind to the patient’s weak A antigens and cause erythrocyte destruction. However, this may only lead to subclinical transfusion reactions. Trauma patients frequently receive group O whole blood, even if they express A or B antigens, without significant haemolysis, provided the donors have low anti-A and anti-B titers (below 100).
A disadvantage of classifying individuals with a weak A subgroup as type O is the restricted range of available transfusion products. For blood group A over 80% of the European donor population is suitable, offering more options compared to blood group O, which has a prevalence of 45%. This limitation is particularly pronounced for rare blood groups and the need for rare donors. The combination of a weak A subgroup with an additional high-prevalence antigen negative requirement is however exceptionally rare. In a reference laboratory setting, performing reverse grouping can still be of use. It provides additional information about the type of antibodies being investigated. Also, in cases of observed weak reactivity upon determination of A and B antigens (1+/2+), it is advisable to perform reverse grouping to understand the underlying cause.
Another consideration is the difficulty of adjusting ABO group results in current Laboratory Information Management systems (LIMS). In cases where a current test result is discrepant with a historical result, and the concluded ABO group has changed, this would involve significant additional work to override an existing ABO group in the LIMS. For patients with acquired B antigens, performing only A and B antigen determination can cause a problem. Acquired B antigens result from bacterial enzymes altering the A antigen to resemble the B antigen. With some reagents, such patients might be classified as blood group AB, although they should receive A group blood. Adjustments in reagents have nearly eliminated detection of acquired B antigens.
After ABO-incompatible organ transplants, passenger lymphocytes can lead to temporary presence of anti-A or anti-B in the circulation, which remain undetected if reverse grouping is not performed. This is particularly crucial in cases of recent organ transplants where patient history might be incomplete.
ABO reverse grouping in donors
For donors who have no ABO group on record, the possible presence of weak or low expression of A or B antigens necessitates anti-A and anti-B antibody determination to ensure correct ABO grouping. Confirming the absence of A antigen through the presence of anti-A antibodies is essential, especially in detecting weak subgroups and chimerism. Thus, omitting antibody determination is undesirable in the initial ABO typing of donors.
Discussion
While there are advantages to omitting ABO reverse grouping, such as reducing unnecessary workload, it may not be prudent at this time. A thorough review over the coming years to identify instances where incorrect conclusions might have been drawn, without antibody determination, can help formulate exceptions and reduce unnecessary procedures. In summary, long-standing practices should be periodically reviewed to assess their continued necessity, and proposed changes should be assessed to ensure streamlining testing procedures does not introduce unnecessary risk to patients.
