BACHELOR OF BIOMEDICAL SCIENCES

BASIC TRANSFUSION SCIENCE AND BLOOD BANKING

BMC 125

CHAPTER 7: ABO DISCREPANCIES

BIOMEDICAL SCIENCE, FACULTY OF MEDICINE

 Learning Outcome
At the end of the lecture, students should able
to
• Explain the discrepancy ABO forward grouping
• Explain the discrepancy of ABO reverse
grouping
• Describe resolving of ABO discrepancy

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 ABO Discrepancy
• Definition: When the results of the forward
grouping (patient cells) do not correspond to
the results of the reverse grouping (patient
serum) or abnormal reactivity is present (i.e.
Mixed Field):
1. Strength of reaction
• Weak or missing
2. Additional reactions
3. Abnormal reactions
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 HINT
• ABO forward and reverse reactions are
typically very strong: 3+ to 4+. Weaker
reactions should immediately send up red flags
indicating that something is wrong.

HINT
• Since production of ABO antigens is genetically
controlled they are less vulnerable to problems
than does the production of ABO antibodies.
Therefore we see more problems in which
grouping: Forward or Reverse?
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 Patient Anti-A Anti-B A1-Cells B-Cells
A 4+ 1+ 0 4+
B 0 4+ 1+ 0
C 4+ 4+ 1+ 0
D 0 3+ 0 0

Patient A: Additional reaction with anti-B and patients cells.

Patient B: Weak reaction with patients serum and A1-cells.
Patient C: Additional reaction with patients serum and
A1-cells.
Patient D: Missing reactions with patients serum A1-cells
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 Problems with The Forward Grouping
Two cell populations
• Mixed Field reactions observed
• Causes: Recent transfusion (O cells to an A
patient), Bone marrow transplant
Testing using Serum or plasma
suspended Patient RBCs
• Can cause non specific aggregation of rbc’s
1. Increased serum proteins: Multiple Myeloma patient
2. Contamination in cord blood sample: Wharton’s jelly
3. Infusion of macromolecular solutions: Dextran, etc.
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Problems with The Forward Grouping
Potent cold reacting autoagglutinins
• Antibody coats patient RBC and agglutinate
spontaneously in the diluent
Weakened Antigen Expression
• Subgroups of A
• Some leukemia’s, Hodgkin's disease
• Excess soluble A and B blood group substances
– Carcinoma of the stomach and/or carcinoma of the
pancreas
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 Problems with The Forward Grouping:

Extra ABO antigens

1. Acquired ‘B’ Antigen
a) Microbial deacetylating enzymes such as E. coli
cleave off the N-Acetyl of the ‘Group A’ N-acetyl-D-
galactosamine immunodominant sugar. The
remaining D-galactosamine becomes similar enough
to the Group B D-galactose immunodominant sugar
that it DOES react with reagent anti-B.

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• A antigen
• B antigen

RBC RBC

Glucose
Glucose

Galactose Galactose

N-acetyl
N-
glucosamine
acetylglucosamine
Galactose Galactose

N-acetyl Galactose
Fuc galactosamine Fucose
ose
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 Problems with The Forward Grouping:
Extra ABO antigens

Polyagglutinable state
• Exposure of ‘crypt’ or buried antigens (T, Tk, etc.)
by inheritance or bacterial enzymes – RBC’s
agglutinate with most sera.

• Exposure of T, Tn and Tk (etc.) antigens.

Antibodies to these antigens are present in
virtually all human antisera. If using human
source anti-A and anti-B these cells will
agglutinate. BIOMEDICAL SCIENCE, FACULTY OF MEDICINE
 Problems with The
Reverse Grouping

1. Plasma or 1. Still has fibrinogen and can

incompletely cause fibrin formation (rbc’s
can be caught up and
clotted serum appears as clumping)

2. Increased serum proteins

2. Non specific red (Multiple Myeloma, etc.),
infusion of high molecular
cell aggregation weight plasma expanders
(eg: dextran)
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 Problems with The
Reverse Grouping
1. Cold reacting
1. Unexpected antibodies such as
antibodies to RBCs anti-M, -N or -I
cell suspension 2. Antibody reacts
with diluent and
2. Antibodies to non-specifically
constituents of the involves red blood
reagent RBC’s diluent cells in reaction
causing clumping.
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Problems with The Reverse Grouping:
Unexpectedly Weakened Antibodies
Immunodeficient due to therapy or disease
– Immunosuppressive drugs
– Certain leukemia’s (CLL) or lymphoma’s
(malignant lymphomas) have
hypogammaglobulinemia
Age related
• Very young: <6 months of age
• Very old: >65 years of age
Dilutional Effect
• Plasma Exchange, Transfusion, etc. dilutes out
patient antibodies
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 Popular LAB CAUSES Of
ABO Discrepancies

1. Poorly labeled specimen OR test tubes

2. Patient RBC suspension too heavy or light
3. Wrong specimen put in Patient’s labeled
test tubes
4. Oh? Is hemolysis really a Pos. Rx’n?
5. Wrong results recorded on Pt. Form
6. Didn’t follow manufacturer’s instructions
7. Poor centrifugation: over or under!
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 Popular LAB CAUSES Of
ABO Discrepancies

Didn’t add:
1. Patient Serum
2. Reagents
3. Correct Reagent
Reaction Reading:
4. Shaking tubes while looking elsewhere
5. Shaking tubes too hard
6. Shaking tubes too gently or not completely re-
suspending cell button
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 ABO Discrepancy
When an ABO Discrepancy is
encountered:
1. Results must be recorded, but interpretation of the
ABO group must be delayed until the discrepancy
is resolved…by you!
2. Begin follow up by getting an accurate patient
history – age, medications, diagnosis, etc.
3. Repeat testing to rule out tech errors such as
mislabeling, adding reagents, wrong patient
sample, etc.
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 Resolving
ABO Discrepancies

1. Repeat testing on 1. From the beginning: re-label

the same sample… tubes, re-drop patient and
reagent drops, etc.

2. Repeat testing using 2. Many labs make the patients

saline suspended red blood cell suspension
and/or washed with the patient’s
patient red blood serum/plasma. If the patient
cell’s. Saline has increased plasma
Replacement. proteins it can cause non-
specific red cell aggregation.
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 Resolving
ABO Discrepancies

1. Weak or missing 1. Incubate test system at

reactions? room temperature for 15-30
minutes! Get patient history.

2. Redraw Patient!!
2. Mislabeled or
a) ALL of the above: any labeling
contaminated error may account for the
specimen: problem and needs to be
redrawn.
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 Resolving
ABO Discrepancies

1. Get patient history.

a) Recent transplant: two cell populations
b) Recent transfusion: two cell populations
and/or dilutional effect
c) Patient medication
d) etc., etc., etc.

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 Resolving
ABO Discrepancies

1. Test patient cells with 1. For suspected

anti-A1 (Dolichos subgroups of A
biflorus), anti-A,B or
anti-H (Ulex europaeus)
2. Ditto!
2. Test patient serum with
A1 or A2 cells

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 Resolving
ABO Discrepancies

1. Review Antibody 1. Can react with

Screening tests reagent A1 and B cells
1. Allo antibody or cold
reactive allo or auto Ab

2. Incubate tests and 2. Should strengthen

controls for 10-30 weakened ABO
minutes room antibody reactivity!
temperature
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 Anti-A Anti-B A1-Cells B-Cells
3+ 0 0 1+

Problem: Reverse grouping - weakened patient

antibody
Causes: Age related (>65, infant),
immunosuppressed or immunocompromised,

Resolution: Incubate Room Temperature 15-30

minutes and respin. Check Patient history.
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 Anti-A Anti-B A1-Cells B-Cells
3+ 1+ 0 4+

Problem: 1+ Reaction with Anti-B. Appears to

have additional antigens.

Causes: Acquired ‘B’ antigen.

Resolution: Patient history – bowel obstruction,

carcinoma of the bowel. (E. coli deacetylation of the
Group A antigen.)
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 Anti-A Anti-B A1-Cells B-Cells
2+ 0 1+ 4+

Problem: Weak forward anti-A and 1+ reaction

with A1 Cells.

Causes: 1)Subgroup of A – A2 with anti-A1.

2) Unexpected cold reacting antibody to antigen on
reagent A1 cells.
Resolution:1) Test patient cells with anti-A1 lectin
and with patient serum test A2 cells
2) Antibody screen should demonstrate unexpected
cold reacting antibody. BIOMEDICAL SCIENCE, FACULTY OF MEDICINE
 REFERENCES
• Denise M. Harmening PhD MLS(ASCP)
CLS(NCA),2012.Modern Blood Banking &
Transfusion Practices 6th Edition.by F.A.Davis
company united sate of America.
• Eva D.Quinley Ms MT
(ASCP)SBB.2010.immunohaematology Principles and
practice,3rd ed( Lippincott Williams and Wikins)
• Jeffrey Mc Cullough.2011.Trnasfusion Medicine 3 rd
ed,wiley Blackwell publication ,United states.
• Harvey G.Klein,David J Anstee 2013,Mollisons blood
transfusion in clinical medicine,12 th ed,Wiley-Blackwell.
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Thank you

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