The blood group systems (ABH)

ABH ANTIGEN

The inheritance of the ABO blood group has demonstrated that each individual inherits one ABO gene from each parent and these two genes determine which Ags are present on RBCs membrane. A Locus termed H and the final product of the genes at that locus, H antigen, is necessary for the expression of normal ABO antigens. ABH Ags of the RBC membrane are found partly as glycolipids, but primarily as glycoproteins. It may also occur in the secretion as glycoproteins. Ags belonging to ABH blood group system are present on RBCs and other body cells and body fluids. The presence of A,B, and O Ags on RBCs depends upon the allelic genes, A,B, and O. An H genes at a separate locus codes for the precursor substance on which the A and B gene products act. The products of the A and B genes are enzymes that act as a specific transferase. H gene products is an enzyme that produce H substance. The O gene is a silent allele. It does not alter the structure of H substance.

ABO Genetics

Genes at three separate loci control the occurrence and location of A and B antigens.

1.Hh genes – H and h alleles

•H allele codes for a fucosyltransferase enzyme that adds a fucose on Type 2 chains (primarily) to form the H antigen onto which A and B antigens are built on red blood cells.

•h allele is a silent allele (amorph). A, B and H antigens are built on oligosaccharide chains of 4 types.

The most common forms are Type 1 and Type 2.

Type 1: #1 carbon of Gal is attached  to the #3 carbon of GlcNAc.

Type 2: #1 carbon of Gal is attached to the #4 carbon of GlcNAc.

Type I and Type II Precursors

•There are two potential precursors substances for ABH antigens Type I and Type II

•Both are comprised of identical sugars but the linkage of the terminal sugars differs in the two types

•Type I precursor has a terminal galactose linked to a subterminal N-acetylgluosamine in a 1-3 linkage

•These same sugars combine in a 1-4 linkage in type II precursor

•ABH Ags on red cells are derived from Type II chains whereas the ABH Ags in plasma are made from both types I & II precursors .

Formation of A, B & H Antigen

The ABO genes do not code for the production of ABO antigens, BUT rather produce specific glycosyl transferases that add sugars to a basic precursor substance on the RBCs. A donor nucleotide derivative supplies the sugar that gives ABO specificity (is the sugar molecule that complete the antigenic determinant when combined with the precursor substance. The inheritance of at least one H gene (HH or Hh) elicits the production of an enzyme called, α-2-L-Fucosyl transferase, which transfers the sugar from the guanosine diphosphate L-fucose (GDP-Fuc) donor nucleotide to the terminal galactose of the precursor chain. The H substance must be formed for the other sugars to be attached in response to an inherited A and /or B genes.

The H antigen is found on the RBCs when there is an Hh or HH genotypes but  NOT with the hh genotype.

The A antigen is found on the RBCs when there is  Hh, HH, and A/A, A/O or A/B genotypes.

The B antigen is found on the RBCs when there is Hh, HH, and B/B, B/O or A/B genotypes.

RBC Precursor Structure

H Antigen

The H gene codes for an enzyme (fucosylytranferase) that adds a Fucose to the terminal sugar of a Precursor Substance (PS*). The biochemical structure below constitutes the H Antigen. (h gene is an amorph.)

Formation of the H antigen

Formation of the A  Antigen

The A gene codes for an enzyme that adds GalNAc (N-Acetyl-D galactosamine) to the terminal sugar of the H Antigen.

Formation of the A antigen

Formation of the B Antigen

Donor Nucleotides & Immundominant Sugars responsible for H, A, and B Ags specificity

Antigen	Immunodominant sugar	Glycosyltransferase	Gene
H	L-fucose	L- fucosyl transferase	H
A	N-acetyl-D-galactoseamine	N acetylgalactosaminyl transferase	A
B	D-galactose	D- galactosyl transferase	B

The O allele

Why do Group O individuals have more H antigen than the other groups?

The O gene is a silent allele. It does not alter the structure of the H substance that means more H antigen sites.

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

Secretor Status

•The secretor gene consists of 2 alleles (Se and se)

•The Se gene is responsible for the expression of the H antigen on glycoprotein structures located in body secretions

•If the Se allele is inherited as SeSe or Sese, the person is called a “secretor”

•80% of the population are secretors

Secretors

•Secretors express soluble forms of the H antigen in secretions that can then be converted to A or B antigens (by the transferases)

•Individuals who inherit the sese gene are called “nonsecretors”

•The se allele is an amorph (nothing expressed)

•sese individuals do not convert antigen precursors to H antigen and has neither soluble H antigen nor soluble A or B antigens in body fluids

Secretor Status Summary

•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.

ABO Subgroups

•ABO subgroups differ in the amount of antigen present on the red blood cell membrane

•Subgroups have less antigen

•Subgroups are the result of less effective enzymes. 

•They are not as efficient in converting H antigens to A or B antigens (fewer antigens are present on the RBC)

•Subgroups of A are more common than subgroups of B.

Subgroups of A

The 2 principal subgroups of A are: 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 and A₂B

Reagent anti-A is a mixture of two Abs ;

anti-A which react with both A1 and A2  cells.

anti-A1 which reacts with A cells but not with A2 cells in simple testing .

Anti-A1-lectin: is another source of anti-A1.

lectins are seed extracts that agglutinate human cells with some degree of specificity.

The seeds of the plant Dolichos biflorus serve as the source of the anti-A1 lectin. This reagent agglutinate A1 or A1B cells but does not agglutinate A2 or A2B cells.

Lectins

•Lectin is a seed extract that has antibody specificity

•Lectins do not contain antibodies, instead they contain proteins that react similar to antibodies

•Used to identify certain types of blood group antigens by binding to the carbohydrate determinant of the antigen, resulting in agglutination

•Other use of Lectin is to investigate red cell polyagglutination

•Some examples

•Dolichos biflorus (binds A1 antigen)

•Ulex europaeus (binds H antigen)

A₂ Phenotype

Why is the A₂ phenotype important?

•A₂ and A₂B individuals may produce an anti-A₁

•This may cause discrepancies when a crossmatch is done (incompatibility)

What’s the difference between the A₁ and A₂ antigen?

•It’s quantitative

•There is fewer A₂ antigen sites compared to the many A₁ antigen sites.

Qualitative difference due to ;

•

•1-8 % of A₂ and  22-35 % of A₂B individuals produce a readily identifiable anti-A₁ in their serum.

•

•Quantitative difference

•A2 cells carry  25 % as many A antigen sites as do A1 cells

•A1 individuals make A antigen.

•A2 individuals produce A antigen.

A₁ versus A₂ Phenotype

	Reactions of patient’s red cells with
Blood Group	Anti-A (from B Sera)	Anti-A1 Lectin
A1	+	+
A2	+	Negative

A₁ has 2 antigens A and A₁

A₂ has only one, A antigen

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

Other A subgroups

There are other additional subgroups of A

•A_int (intermediate), A₃, A_x, A_m, A_end, A_el, A_bantu

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, B_x, B_m, and B_el

Amount of H Antigen According to Blood Group

Other ABO conditions

Bombay Phenotype (O_h)

•Inheritance of hh

•The h gene is an amorph and results in little or no production of L-fucosyltransferase

•Originally found in Bombay (now Mumbai)

•Very rare

Bombay

•The hh causes NO H antigen to be produced

•Results in RBCs with no H, A, or B antigen (patient types as 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

•What ABO blood group would you use to transfuse this patient??

ANSWER: Another Bombay

•Group O RBCs cannot be given because they still have the H antigen

•You have to transfuse the patient with blood that contains NO H antigen.

ABO Antibodies

Landsteiner’s Rule

Normal, healthy individuals possess ABO antibodies to the ABO antigen absent from their RBCs

•Most blood group systems (ABO and others) are made up of:

An antigen on a red cell and the absence of it’s corresponding antibody in the serum (if you’re A, you don’t have anti-A)

•If you do NOT have a particular antigen on your red cells then it is possible (when exposed to foreign RBCs) to illicit an immune response that results in the production of the antibody specific for the missing antigen.

RBC Phenotype (antigens)	Frequency (%)	Serum Ab
A	43	Anti-B
B	9	Anti-A
AB	4	——–
O	44	Anti-A,B

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.

•IgM is the predominant antibody in Group A and Group B individuals

•Anti-A

•Anti-B

•React best at room temperature (22-24^oC) or below in vitro.

•Activates complement to completion at 37^oC

• Can cause acute Hemolytic Transfusion reactions

•IgG (with some IgM) is the predominant antibody in Group O individuals

•Anti-A,B (with some anti-A and anti-B)

•RBC Immune form: Predominantly IgG

Time of appearance:

•Usually present within the first 3-6 months of life

•Stable by ages 5-6 years

•Decline in older age (>65 years of age) & in hypogammaglobulinemia

•Newborns may passively acquire maternal antibodies (IgG crosses placenta)

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