The document discusses the ABO blood group system, including its discovery, genetics, biochemistry, antigens, antibodies, and implications for transfusion. Some key points: - Karl Landsteiner discovered the main ABO blood groups (A, B, AB, O) in 1900. The ABO blood type is determined by alleles at a single gene locus. - The antigens are carbohydrate structures on red blood cells. People naturally produce antibodies against antigens they lack. - ABO typing must be accurate to avoid transfusion reactions. Discrepancies can occur due to weak subgroups, diseases, or test issues. Resolving discrepancies helps ensure patient and donor safety.

1. ABO blood group system
Dr R Amita
Assistant Professor
Dept of Transfusion Medicine

2. Discovery
• Karl Landsteiner(1900) discovered human A,B,O groups.
• Von Decastello and Sturli (1902) discovered AB blood group.
• Von Dungern and Hirszfeld(1911) divided group A into 2 subgroups
A1 and A2.
ABO system is classified into 6 groups: A1, A2, A1B, A2B, B, AB and O

3. Landsteiner’s law
• 1.If an agglutinogen is present on red blood cell membrane ,the
corresponding agglutinin must be absent in the plasma.
• 2.If an agglutinogen is absent on red blood cell membrane, then
corresponding agglutinin must be present in the plasma.

5. Antigens
• Appear in the sixth week of fetal life.
• Present on red cell membrane, WBC, platelets and in other tissues
like salivary glands, pancreas, kidney, body fluids
• Exception CNS

6. ABO gene
• The ABO blood type is controlled by a single gene (the ABO gene)
with three types of alleles i, IA, and IB.
• The gene encodes a glycosyltransferase
• The gene is located on the long arm of the ninth chromosome
(9q34).
• IA allele gives type A, IB gives type B, i gives type O.
• Co dominant
• O group : Only ii AB group : IAIB
• A group : IAIA or IAi B group : IBIB or IBi

7. Genetics
• Inheritance of genes follows Mendelian Law
• Bernstein theory: there is one locus on a chromosome at which
any of the three alleles may be located

8. ABO Antigen Genetics
• The presence or absence of the ABH antigens on the red blood cell
membrane is controlled by the H gene (chr 19)
• The presence or absence of the ABH antigens in secretions is
indirectly controlled by the Se gene (chr 19)
• H gene – H and h alleles (h is an amorph)
• Se gene – Se and se alleles (se is an amorph)
• ABO genes – A, B and O alleles

9. Biochemistry
• Precursor: Paragloboside/Glycan
• Type I precursor : terminal galactose linked to a
subterminal N-acetylgluosamine in a 1-3 linkage.
• Type II precursor : same sugars combine in a 1-4 linkage
• ABH antigens on RBC are derived from Type II chains
• Blood group substances in secretion are made from
both types I & II precursors

10. Precursor Substance
Glucose
Galactose
N-acetylglucosamine
Galactose
Precursor
Substance
(stays the
same)
RBC

11. H substance
• H gene (FUT1 gene) leads to
production of an enzyme
α-2-L-Fucosyl transferase,
which transfers fucose to the
terminal galactose of the
precursor
Glucose
Galactose
N-acetylglucosamine
Galactose
RBC
Fucose

12. A antigen
• The “A” gene codes for
N-acetylgalactosaminyltransferase
that adds N-acetylgalactosamine to
the terminal sugar of the H antigen
Glucose
Galactose
N-acetylglucosamine
Galactose
RBC
Fucose N-acetylgalactosamine

13. B antigen
• The “B” gene codes for
D-galactosyltransferase
that adds D-galactose to the
terminal sugar of the H
antigen
Glucose
Galactose
N-acetylglucosamine
Galactose
RBC
Fucose Galactose

14. ABO Subgroups
• ABO subgroups differ in the amount of antigen present
on the red blood cell membrane
• Subgroups have fewer antigens are present on the RBC
• Subgroups are the result of less effective enzymes (not
as efficient in converting H antigens to A or B antigens)
• Subgroups of A are more common than subgroups of B

15. Subgroups of A
• A1 and A2
• Both react strongly with reagent anti-A
• To distinguish A1 from A2 red cells, the lectin Dolichos
biflorus is used (anti-A1)
• 80% of group A or AB individuals are subgroup A1
• 20% are A2 or A2B

16. A2 Phenotype
• The A2 gene doesn’t convert the H3 & H4 to A very well
• The result is fewer A2 antigen sites compared to the
many A1 antigen sites.
• A2 and A2B individuals may produce an anti-A1
• This may cause discrepancies when a crossmatch is
done.

17. A1 and A2 Subgroups
Anti-A
antisera
Anti-A1
antisera
Anti-H
lectin
ABO
antibodies
in serum
# of
antigen
sites per
RBC
A1
4+ 4+ 0 Anti-B 900 x103
A2
4+ 0 3+ Anti-B &
anti-A1
250 x103

18. Other A subgroups
• There are other additional subgroups of A
• Aint (intermediate), A3, Ax, Am, Aend, Ael, Abantu
• A3 red cells cause mixed field agglutination when
polyclonal anti-A or anti-A,B is used
• Mixed field agglutination appears as small
agglutinates with a background of unagglutinated RBCs
• They may contain anti-A1

21. B Subgroups
• B subgroups occur less than A subgroups
• B subgroups are differentiated by the type of reaction
with anti-B, anti-A,B, and anti-H
• B3, Bx, Bm, and Bel

22. Other ABO conditions
• Bombay Phenotype (Oh)
• Inheritance of hh
• Missense mutation of FUT1 gene
• The h gene is an amorph and results in little or no
production of L fucosyltransferase
• Originally found in Bombay (now Mumbai) by Bhende in
1952
• Very rare (Frequency in India 1:10000)

23. Bombay group
• The hh causes NO H antigen to be produced
• Results in RBCs with no H, A, or B antigen (Cell group: O)
• Bombay RBCs are NOT agglutinated with anti-A, anti-B, or anti-H
(no antigens present)
• Bombay serum has strong anti-A, anti-B and anti-H, agglutinating
ALL ABO blood groups.
• Group O RBCs cannot be given because they still have the H
antigen
• Transfuse the patient with blood that contains NO H antigen

24. Parabombay phenotype
• H antigen is weakly expressed on RBCs.
Weak expression of A, B, H antigens on the red cells (Due
to passive adsorption from secretion) which react weakly
with antisera to A, B, H antigens
• H antigen is present in the secretions, but there is no
expression on red cells. Serum contains anti-H antibodies
• Genotype:hh/Sese or hh/SeSe

26. ABO Antigens in Secretions
• Secretions include body fluids like plasma, saliva, synovial fluid,
etc
• Blood Group Substances are soluble antigens (A, B, and H) that
can be found in the secretions.
• This is controlled by the H and Se genes
• Se gene (FUT2gene) encodes α2 L fucosyltransferase which
modifies type 1 precursor to H substance
• If the Se allele is inherited as SeSe or Sese, the person is called a
“secretor”
• 80% of the population are secretors

27. Secretor Status
• The Se gene codes for the presence of the H antigen in secretions,
therefore the presence of A and/or B antigens in the secretions is
contingent on the inheritance of the Se gene and the H gene
Se gene (SeSe
or Sese)
H antigen in
secretions
A antigen
B antigen
se gene (sese) No antigens secreted
in saliva or other body
fluids
and/or

28. ABO Group
ABH
Substances
Secretors (SeSe or Sese): A B H
A +++ 0 +
B 0 +++ +
O 0 0 +++
AB +++ +++ +
Non-secretors (sese):
A, B, O, and AB 0 0 0
Sese + h/h (no H antigen)  no antigens in secretions

29. H antigen
• Certain blood types possess more H antigen than others:
• The O gene is a silent allele. It does not alter the structure of the
H substance….that means more H antigen sites
O>A2>B>A2B>A1>A1B
Greatest
amount of H
Least
amount of H

30. Group O Group A
Many H
antigen sites
Most of the H antigen sites in a
Group A individual have been
converted to the A antigen
Fewer
H antigen
sites
A
A A
AA
Group O Group A

31. ABO antibodies
• Natural antibodies: does NOT require the presence of a
foreign red blood cell for the production of ABO
antibodies.
• ABO antibodies are “non-red blood cell stimulated”
probably from environmental exposure.
• Titer of ABO Abs is often reduced in elderly and in
patients with hypogammaglobulinemia and infants (until
3 -6 months of age)

32. ABO antibodies
• IgM is the predominant antibody in Group A and Group B
individuals
• Anti-A
• Anti-B
• IgG (with some IgM) is the predominant antibody in
Group O individuals
• Anti-A,B (with some anti-A and anti-B)

33. Anti-A
• Group O and B individuals contain anti-A in their serum
• However, the anti-A can be separated into different
components: anti-A and anti-A1
• Anti-A1 only agglutinates the A1 antigen, not the A2
antigen
• Occurs in 1-8% of A2 and 22-30% of A2B
• There is no anti-A2.

34. Anti-A,B
• Found in the serum of group O individuals
• Reacts with A, B, and AB cells
• Predominately IgG, with small portions being IgM
• Anti-A,B is one antibody, it is not a mixture of anti-A and
anti-B antibodies

35. Anti H antibody
• A1 gene very efficiently converts all of H substance to A
antigen,
• Therefore some A1 and A1B individuals may have anti H
• IgM cold agglutinin
• Best reacts at room temperature

36. Anti-H
Auto-Anti-H
Clinically
Significant
No
Abs class
IgM
Thermal range
4 - 15
HDNB
No
Transfusion Reactions
Extravascular Intravascular
No yes
Allo-Anti-H (Bombay group)
Clinically
Significant
Yes
Abs class
IgM, IgG
Thermal range
4 - 37
HDNB
Yes
Transfusion Reactions
Extravascular Intravascular
Yes Yes

37. Frequency of blood group system
RBC Phenotype Frequency (%) Serum Ab
A 24 Anti-B
B 30 Anti-A
AB 6 --------
O 40 Anti-A,B

38. Transfusion

39. ABO discrepancies
• Group I Discrepancies -
• These are associated with unexpected reactions in the reverse
grouping due to weakly reacting or missing antibodies.
• Includes:
• Infants less than 4-6 month of age
• Elderly patients
• Severe hypogammaglobulinemia
• ABO incompatible HPC transplantation

40. • Resolution:
• Enhancing weak or missing reaction by incubating the patient’s
serum with reagent A1 and B cells at room temperature for 15-30
minutes
• Serum cell mixture is incubated at 4⁰C for 15-30 minutes
• An autocontrol and O cell control must always be tested
concurrently to detect reactivity of other commonly occurring
cold agglutinins eg: anti I

41. • Group II discrepancies
• These are associated with unexpected reactions in forward grouping due to
weakly reacting or missing antigen
• Includes:
• Weak subgroups of A or B
• Weakening of A or B antigen in malignancies
• Acquired B phenotypes:
• results from the action of bacterial deacetylase, which converts N-
acetylgalactosamine to ẞ-galactosamine, which is very similar to galactose, the
chief determinant of B.
• ‘passenger antigen’ type is caused by adsorption of B-like bacterial products on
to O or A cells but occurs only in vitro.
• Out of group transfusion or ABO mismatched HPSCT
• Neutralization of anti A and anti B typing reagent by high concentration of A or B
soluble substances in serum with serum or plasma suspended red cell

42. • Resolution of group II discrepancies
• Weaker reactions with antisera can be resolved by enhancing reaction of
antigen with respective antisera by incubating test mixture at room
temperature for 15-30 minutes
• Sub groups causing group discrepancies can be resolved by adsorption
elution studies
• Acquired B phenomenon can be resolved by lowering PH of monoclonal
antisera. Anti B in the serum of acquired B person does not agglutinate
autologous red cells (autocontrol negative).
• Secretor status of person can resolve acquired B, saliva of acquired B
person contains A substance not B substance.
• High concentration of A or B substance causing group discrepancies can
be resolved by saline washing of red cells

43. • Group III discrepancies
• These are associated with protein or plasma abnormalities, rouleaux formation
and pseudoagglutination.
• Includes
• Elevated level of globulin from e.g. multiple myeloma, waldenstorm
macroglobulinemia, Hodgkin lymphoma.
• Elevated level of fibrinogen.
• Small fibrin clot in plasma or incompletely clotted serum can be mistaken for
red cell agglutinates in reverse grouping.
• Sample with abnormal concentration of serum proteins, altered serum protein
ratio, or high molecular weight volume expanders can aggregate reagent red
cells and can mimic agglutination.
• Rouleaux will disperse when suspended in saline. True agglutination is stable in
the presence of saline

44. • Group IV discrepancies are due to miscellaneous problems.
• Recent transfusion of out of group plasma containing component.
• Cold alloantibodies (e.g. anti M) or autoantibodies (e.g. anti I), pH
dependent autoantibodies, a reagent dependent antibody (e.g.
EDTA, paraben) leading to unexpected positive eaction.
• Recent infusion of IvIg which can contain ABO isoagglutinins.
• Mixed field agglutination with circulating red cell of more than
one ABO type.

45. Resolving ABO discrepancy