ABO Discrepancies
Student Name: Ahmed Refaei
Student Number:
Institutional Affiliation: Jazan University
Date: 10/5/2025
Abstract
The foundation of transfusion medicine is the ABO blood group system,
where accurate blood typing is necessary to prevent hemolytic transfusion
responses. The interpretation of blood type findings can occasionally be
complicated by variations between forward (antigen) and reverse (antibody)
typing, known as ABO discrepancies. A wide range of factors, including abnormal
serum proteins, low or missing naturally occurring antibodies, inadequate antigen
expression, and uncommon genetic and pathologic diseases, might contribute to
these disparities. Based on their etiologic cause, ABO incompatibilities can be
divided into four broad types, each of which requires a unique investigative and
remedial procedure. The classification, causes, and resolution of ABO
discrepancies are critically reviewed in this paper, with a focus on their clinical
significance and the paramount need of precise laboratory methods in
maintaining transfusion safety and diagnostic consistency.
Introduction
Karl Landsteiner first described the ABO blood type system in 1901, and it
serves as the basis for both immunohematology and transfusion medicine.
Because it contains naturally occurring antibodies that might react with
incompatible antigens and cause potentially catastrophic hemolytic transfusion
events, it is considered clinically relevant. Therefore, a crucial first step in
ensuring the safety of organ transplants, blood transfusions, and other medical
operations involving blood compatibility is accurately determining a patient's ABO
blood group.
ABO typing consists of two complimentary tests: reverse typing, which uses
reagent red blood cells to find naturally occurring anti-A and/or anti-B antibodies
in the patient's serum, and forward typing, which uses known antisera to find the
presence of A and/or B antigens on the patient's red blood cells. Forward and
reverse typing findings should ideally match, providing highly reliable
confirmation of the person's ABO group. The so-called ABO discrepancy, however,
occurs when these results do not accord with one another.
Despite being relatively rare, ABO differences provide a significant
challenge to transfusion services. They can occur for a variety of causes, such as
the existence of rare plasma proteins, illness states like leukemia or
immunodeficiency conditions, weak or absent antigens or antibodies, or
uncommon genetic variations like subgroups of A or B. Recurring variances may
also result from technical or administrative mistakes made when handling or
testing specimens. Based on their genesis, they are divided into four major types,
each of which needs a specific research path to be resolved.
To prevent unnecessary transfusions, diagnostic errors, and patient care
delays, ABO inconsistencies must be identified and resolved. The underlying
cause, appropriate troubleshooting techniques, and clinical consequences of ABO
differences must be thoroughly understood by laboratory staff. Examining the
categorization, cause, and resolution of ABO differences while highlighting their
importance in clinical laboratory settings and patient safety is the aim of this
research.
Body
1. Types of ABO Discrepancies
ABO discrepancies are usually divided into four main types. These are based
on whether the problem is with the antigens (on red blood cells), antibodies (in
the plasma), or something else affecting the test results. Knowing the type helps
lab staff figure out what’s going wrong and how to fix it.
Group I: Missing or Weak Antibodies
This is the most common type. It happens when the body doesn’t make
enough antibodies to show a strong reaction in reverse typing. This can
happen in:
o Newborns (their immune systems are still developing)
o Elderly people
o People with weak immune systems (like cancer patients)
o After a recent transfusion
A rare condition known as "chimerism" occurs when a person's body
contains two sets of red blood cells with distinct blood types. There are no
antibodies against either of the red cell antigens in the serum since testing
identifies them as self-antigens. This situation results in an ABO discrepancy,
which frequently happens in fraternal twins, with mixed field reactions in forward
grouping reactions and absent reactions in reverse grouping. More infrequently,
mosaicism causes chimerism in people who are not fraternal twins.
Most group I reactions have an underlying cause that is determined by the
patient's history. In order to verify this disparity, physicians need to strengthen
the weak or absent antibody by allowing the serum to incubate with reagent red
blood cells at room temperature for 15 to 30 minutes. Usually, this results in a
satisfactory reaction. In certain cases, centrifuging the sample is necessary to
bring the cells and antibodies closer together. The material is subsequently
incubated by the clinicians, and a suitable reaction ought to ensue. Clinicians
incubate a mixture of the patient's cells and the reagent cells at 4 °C for 30
minutes if a reaction is still not visible.
Group II: Missing or Weak Antigens
In the forward grouping, patients with group II differences react in an
unexpected way. These differences arise in patients with weakly responding
antigens because of either a low antigen burden on the RBC surface, the existence
of an antigen on the RBC surface that is similar to AB antigens and reacts with the
antibody, or partial cell surface antigen deletion. In forward type, these antigens
provide a weaker-than-expected reaction that is not matched in reverse typing.
This may happen due to:
o Certain types of leukemia
o A or B subgroups (weaker versions of the regular A or B blood types)
o After a bone marrow transplant
o Pregnancy
o Fetomaternal hemorrhage
By increasing the incubation period to up to 30 minutes at room
temperature, clinicians can improve the weak reaction in forward typing. The
combination should be incubated at 4 °C for up to 30 minutes if further
incubation does not result in enhancement. Examining a patient's medical history
can reveal crucial information about what caused the disparity.
Group III: Abnormal Plasma Proteins
Because of protein or plasma abnormalities, patients with group III
discrepancies show a mismatched reactivity between the forward and reverse
categorization. These anomalies cause pseudo-agglutination or rouleaux
development, which produces inaccurate or incomprehensible results. For
example:
o Hypergammaglobulinemia
o Hyperfibrinogenaemia
o Plasma expanders such as dextran or hydroxyethyl starch
o Wharton jelly contamination of cord blood
Agglutination is used in ABO typing to identify antigens and antibodies.
Rouleaux formation can produce false-positive results because it closely mimics
the agglutination of red blood cells. In order to eliminate extra antibodies and
proteins that cause undesired RBC stacking, technicians should wash the sample
with regular saline when rouleaux formation is expected. Usually, the
disagreement is resolved by repeating the test using cleaned cells.
Clinicians may use a saline replacement approach when reverse typing
differences occur. Using this approach, the patient's blood is centrifuged, the
plasma is extracted, and an equal volume of saline is added. The test is then
repeated. Rouleaux development is caused by excess protein in the patient's
serum, which is eliminated by this procedure. After that, the test can be run
without protein interference, producing a result that can be understood.
Group IV: Other Causes (Rare or Unusual)
Agglutination reactions happen without the need for a particular reagent
antibody, and group IV discrepancies are mismatched reactions between the
forward and reverse grouping caused by many factors, such as:
o Cold-reactive autoantibodies
o ABO isoagglutinin
o Non-ABO alloantibodies
o Recent intravenous immunoglobulin infusion
o Reaction to rare antigens or antibodies in the reagent used
Clinicians should take cold autoantibodies in the patient's serum into
consideration when both forward and reverse grouping results from ABO type
show inexplicable positive results. A positive direct Coombs test result is possible.
In order to inactivate immunoglobulin M (IgM) antibodies, clinicians should
incubate the patient's red blood cells at 37 °C. Before doing the test again, the
sample needs to be rinsed three times in saline at 37 °C. This procedure aids in
removing the antibodies' impact on the outcomes. Rarely, 0.01 M dithiothreitol
might be given to the patient's red blood cells to stop IgM antibody agglutination
if the earlier procedures do not work.
Since this usually prevents the autoantibodies from interfering, the test
should be redone at 37 °C if the disagreement occurs in the reverse grouping test.
A cold autoabsorption test ought to be carried out if an unexpected outcome
continues. By incubating the patient's red blood cells with their serum,
autoantibodies react with the patient's RBCs, lowering the concentration of these
antibodies in the serum. This process is known as autoabsorption. Autoantibody-
coated RBCs are removed from the sample by centrifugation prior to testing, and
the residual serum is utilized for reverse blood grouping.
Finding the source of the disparity in this group can be aided by examining
the patient's medical history and current state of health. Clinicians should repeat
the test using a different lot number of the reagent if it is suspected that the
reagent is the cause of the result.
2. What Causes These Discrepancies?
Many things can lead to an ABO discrepancy. Some are natural, like age or
weak immune systems. Others are due to medical conditions like cancer or recent
treatments like transfusions or transplants. Even simple errors—like mislabeled
tubes or contaminated samples—can cause results to appear inconsistent. That’s
why labs always double-check any unusual findings.
Miscellaneous problems may cause ABO discrepancies, including:
Cold reacting autoantibodies that result in spontaneous RBC agglutination
or a positive direct antiglobulin test (DAT)
Mixed red blood cell populations due to massive transfusion
Unexpected alloantibodies unrelated to ABO antigens
Unexpected ABO isoagglutinins
3. Resolution Strategies
Identifying and resolving ABO discrepancies involves a stepwise approach
combining laboratory techniques and clinical correlation. Key strategies include:
Repetition of ABO testing using fresh reagents and careful attention to
technique to rule out technical or clerical errors.
Extended incubation at room temperature or 4°C to enhance weak
antibody reactions, particularly useful in resolving Group I discrepancies.
Use of enzyme-treated red cells or adsorption/elution studies to detect
weak antigens in Group II cases.
Saline replacement technique to eliminate rouleaux and clarify reactions in
Group III discrepancies.
Advanced testing methods, such as:
o Molecular genotyping to detect ABO subgroups or chimerism.
o Direct antiglobulin test (DAT) to identify antibody-coated red cells.
o Patient history review, including recent transfusions, medications, or
diagnoses, to provide clinical context.
Resolution must be guided by both laboratory findings and a detailed
understanding of the patient’s medical condition. In complex cases, consultation
with a transfusion medicine specialist may be warranted.
Conclusion
Although often overshadowed by the predominance of the ABO and Rh(D)
systems, uncommon blood group systems such as Kell, Duffy, Kidd, MNS, and
Lutheran play a vital role in transfusion safety and immunohematologic risk
management. Their clinical relevance is particularly apparent in sensitized
patients, those with unique antigenic profiles, and pregnant women at risk for
alloimmunization. They can harbor highly immunogenic antigens capable of
triggering adverse transfusion reactions and complicating perinatal management.
Exclusions from routine serologic screening point to the need for broader
incorporation of molecular genotyping and newer immunohematology techniques
in finding and treating these antibodies. As more progress is made with
personalized medicine and precision transfusions, a more profound
understanding and integration of these underused systems into routine
procedures will be ever more critical. Eventually, improved awareness, larger rare
donor databases, and improving diagnostic capacity will collectively increase
patient safety and outcomes from transfusions across a broad range of clinical
situations.
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