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ABO subgroups and discrepancies

17 min

  • Use reverse grouping, lectin tests, and history to explain a weak ABO forward reaction
  • Assign an adult ABO group from concordant forward and reverse results
  • Choose the next check for a discrepant ABO reaction pattern

Read the full reference

Try first

Try first

A 2-day-old's red cells react 3+ with anti-A, 0 with anti-B and 0 with anti-A₁ lectin from Dolichos biflorus. How is the subgroup reported?

Right. The next section explains why.

The next section explains it.

The next section explains it.

The next section explains it.

Get the idea

Forward and reverse must agree

Forward grouping tests the patient's red cells with anti-A and anti-B. Reverse grouping tests the plasma with A1 and B reagent cells. CLIA requires both, and the two must support one group before the laboratory records it.1,3

ABO groupAnti-AAnti-BA1 cellsB cells
ONegativeNegativePositivePositive
APositiveNegativeNegativePositive
BNegativePositivePositiveNegative
ABPositivePositiveNegativeNegative

A young infant's expected antibodies are unreliable, so the infant procedure generally assigns the group from forward grouping.3

Find the side that is off

When the two sides disagree, the weakest or unexpected reaction usually marks the affected side. Identity, labeling, reagents and technique are checked first.3 Then the pattern points to the next check:2,3

  • A weak or missing reverse reaction fits a young infant, low immunoglobulin or immunosuppression. The reverse grouping is extended under the procedure, and the history is reviewed.
  • A weak or mixed-field forward reaction fits a subgroup, a recent transfusion or a transplant. The history comes first.
  • An extra reverse reaction fits anti-A1, a cold antibody or rouleaux. Group O cells, A2 cells, an autocontrol and a look under the microscope separate them. Saline replacement disperses rouleaux.

The group is recorded only after the discrepancy is resolved. Red cells needed before then are group O.2

A subgroup needs more than one reaction

A2 cells react with anti-A and stay nonreactive with the anti-A1 lectin from Dolichos biflorus. Some people with A2 make anti-A1, usually a cold antibody that shows as an extra reaction with A1 reagent cells. A newborn's weaker A expression can also leave the lectin nonreactive.3 Reverse grouping, lectin tests and the history are read together before a subgroup is named.3

References
  1. Standard: Immunohematology, 42 CFR §493.1271 (2026). Accessed September 27, 2026. https://www.ecfr.gov/current/title-42/chapter-IV/subchapter-G/part-493/subpart-K/section-493.1271
  2. Association for the Advancement of Blood & Biotherapies. Fundamental Standards for Blood Collection and Transfusion. 2nd ed. AABB; 2025. Effective January 1, 2026. Accessed September 27, 2026.
  3. Bloch EM, Campbell-Lee S, McKenna DH Jr, Montemayor-Garcia C, Schwartz J, Shaz B, Storry J, eds. Technical Manual. 22nd ed. AABB; 2026.

Watch one

Walter Brinkerhoff, 84, a retired ferry captain with chronic lymphocytic leukemia and low immunoglobulin levels, has a type and screen before hip repair. His record from 2 years ago shows group A.

TestReaction
Anti-A, patient cells4+
Anti-B, patient cells0
A1 cells, patient plasma0
B cells, patient plasma0

What is the next check?

  1. Check the specimen label, the request, the reagents and the recorded reactions. All are in order.

    A wrong specimen or a missed reagent explains any pattern, so identity and technique come first.

  2. The forward grouping reads group A. The reverse grouping, with no anti-B, reads group AB. The results disagree.

    The two sides must support one group.

  3. The missing reaction is anti-B, so the reverse grouping is the side that is off.

    The weak or missing reaction marks the affected side.

  4. His age, his leukemia and his low immunoglobulin all fit weak antibody production, and his record agrees with the forward grouping.

    History often explains a missing plasma antibody.

  5. Repeat the reverse grouping with extended room-temperature incubation under the procedure, with group O cells and an autocontrol.

    Longer incubation lets weak antibody react, and the controls show whether the plasma reacts with every cell.

  6. Record the group only when the reverse grouping supports it. Red cells needed before then are group O.

    A group recorded from one side can be wrong.

The missing anti-B points to low antibody production. The next check is an extended reverse grouping with group O cells and an autocontrol, and the ABO group waits for its result.

Your turn

Problem 1 of 3

An adult’s cells react with anti-A only. Plasma reacts with B cells only. What is the ABO group?

Incorrect. Group O cells lack both A and B antigens, and expected plasma reacts with both A₁ and B cells.

Correct. The cells express A antigen and the plasma contains expected anti-B.

Incorrect. Group B gives the opposite pattern: anti-B reactivity and expected anti-A in plasma.

Hint
  1. Forward grouping reads the antigens on the patient's cells.
  2. The plasma holds antibody to the antigen the cells lack.

Review D typing

Problem 2 of 3

A 67-year-old with multiple myeloma has these results. Under the microscope, the cells in the A1 cell tube stack like coins. TestReactionAnti-A, patient cells4+Anti-B, patient cells0A1 cells, patient plasma2+B cells, patient plasma4+O cells, patient plasma2+Autocontrol2+ What is the next check?

The reverse grouping does not yet support group A. The discrepancy is resolved before a group is recorded.

Assigned ABO from one side of a discrepancy

A weak subgroup, a mixed population after transfusion or transplant, or a missing plasma reaction can each make one side of ABO testing misleading. A group assigned from that side can be wrong and lead to incompatible components, which is why the discrepancy is resolved before assignment. Red cells needed urgently in the meantime are group O.

Anti-A₁ reacts with A₁ cells and spares group O cells and the patient's own cells. Here the O cells and the autocontrol react too.

Coin stacks with reactive O cells and autocontrol point to rouleaux from the myeloma protein. Saline replacement disperses rouleaux and leaves true agglutination in place.

Hint
  1. Look at the group O cells and the autocontrol.
  2. Ask what in myeloma plasma can make cells stick together without antibody.

Review ABO discrepancies

Problem 3 of 3

A 58-year-old man's record from last year shows group A. Five days ago he received 4 units of group O red cells at another hospital. Today his cells react 2+ mixed field with anti-A and 0 with anti-B. His plasma reacts 0 with A1 cells and 4+ with B cells. What explains the weak forward reaction?

A mixed field has several causes. His transfusion history explains it before any subgroup is considered.

Assigned a subgroup from one weak forward reaction

A weak forward reaction can come from an A subgroup, a mixed red-cell population, immature antigen expression in a neonate, or reagent and method differences. Naming a subgroup from that reaction skips the reverse grouping, lectin testing, and history that separate these causes, and a single neonatal lectin result leaves the subgroup unresolved.

Donor O cells stay unagglutinated beside his A cells, which gives a mixed field. His reverse grouping and his record both support group A.

A reagent problem would weaken every reaction evenly. A mixed field shows two cell populations.

The lectin would test a mixture of donor and patient cells. His transfusion history explains the reaction.

Review A1, A2, and weaker expression

Use it

  • Evangeline Marsh, 52, a beekeeper, has a type and screen before planned surgery.
  • She has not been transfused or pregnant in the past 3 months, and she has no previous record at this hospital.
  • The antibody screen is negative.
Decision 1 of 3

What kind of discrepancy is this, and what is the next check? TestReactionAnti-A, patient cells4+Anti-B, patient cells0A1 cells, patient plasma1+B cells, patient plasma4+O cells, patient plasma0Autocontrol0

Only the A₁ cells react unexpectedly, with O cells and the autocontrol nonreactive. The lectin and A₂ cells test for A₂ with anti-A₁.

Group A plasma should leave A₁ cells unagglutinated. The 1+ reaction is resolved before a group is recorded.

Assigned ABO from one side of a discrepancy

A weak subgroup, a mixed population after transfusion or transplant, or a missing plasma reaction can each make one side of ABO testing misleading. A group assigned from that side can be wrong and lead to incompatible components, which is why the discrepancy is resolved before assignment. Red cells needed urgently in the meantime are group O.

Rouleaux would also make O cells and the autocontrol react. Both are nonreactive.

Review ABO discrepancies

Decision 2 of 3

Surgery moves up before the lectin result is back, and red cells are requested. Which ABO group is issued?

Her group is not recorded until the discrepancy is resolved. A group taken from one side can be wrong.

Assigned ABO from one side of a discrepancy

A weak subgroup, a mixed population after transfusion or transplant, or a missing plasma reaction can each make one side of ABO testing misleading. A group assigned from that side can be wrong and lead to incompatible components, which is why the discrepancy is resolved before assignment. Red cells needed urgently in the meantime are group O.

AB red cells carry A antigen and B antigen, and her plasma contains anti-B. They are incompatible.

Red cells needed before an ABO discrepancy is resolved are group O. The investigation continues.

Review ABO discrepancies

Decision 3 of 3

Her cells are nonreactive with anti-A₁ lectin, and her plasma is nonreactive with A₂ cells. What does the pattern show?

An autoantibody would react with her own cells, and the autocontrol is nonreactive.

Her A cells lack A₁, and her plasma reacts with A₁ cells and spares A₂ and O cells. Forward grouping, lectin, selected cells and history agree on A₂ with anti-A₁.

Acquired B shows as an unexpected reaction of the patient's cells with anti-B. Her cells are nonreactive with anti-B, and the extra reaction sits on the plasma side.

Review A1, A2, and weaker expression

The clue that settled this case is the reaction with A1 cells alone, with group O cells and the autocontrol nonreactive. That pattern pointed to anti-A1, and the lectin and A2 cells confirmed an A2 phenotype. Until then, red cells were group O.

Keep

Sources checked