Chapter 06 The ABO Blood Group System

The ABO Blood Group System

• The ABO system is crucial in transfusion medicine because individuals possess antibodies against ABO antigens absent from their red blood cells without prior exposure.
• Incompatible ABO transfusions can lead to immediate red cell lysis and severe reactions.
• Detecting ABO incompatibility is the basis for pre-transfusion testing.
• Even now ABO incompatibility is a leading cause of hemolytic transfusion reaction fatalities.

Transfusion-Related Fatalities (FY2009)

• TRALI was the most frequent cause of death in FY 2009.
• There were four fatal hemolytic transfusion reactions, because of ABO incompatibility.

Historical Perspective

• Karl Landsteiner discovered the ABO system, marking the beginning of understanding individual uniqueness based on RBC antigens.
• Landsteiner performed forward and reverse grouping in 1901.
• Forward grouping uses known antisera (anti-A, anti-B) to detect antigens on RBCs.
• Reverse grouping detects ABO antibodies in patient serum using known reagent RBCs (A1 and B cells).
• ABO forward and reverse grouping tests are essential for all donors and patients.
• ABO grouping is the most frequent blood bank test.
• An inverse relationship is always found between forward and reverse typing, that serves as a check.
• Individuals with A antigens on red cells have anti-B in their serum.
• Bacteria, pollen, and other substances stimulate anti-A and anti-B production.
• The regular occurrence of anti-A and/or anti-B confirms ABO grouping results.

ABO Blood Group Frequencies

• Group O and A are most common, AB is the rarest.
• Group B is more frequent in blacks and Asians than in whites.
• Group A is less common in blacks and Asians compared to whites.
• Subgroup A2 is rarely found in Asians.

ABO Antibodies

• Individuals produce antibodies against absent A and/or B antigens and are naturally occurring.
• ABO antibodies are primarily IgM that activate complement and react at room temperature or colder.
• ABO antibodies cause strong direct agglutination during testing.
• Antibody production starts at birth, detectable at age 3 to 6 months.
• Most antibodies in cord blood serum are maternal, making serum ABO testing invalid for newborns.
• Only forward grouping is performed on cord blood from newborn infants.
• Antibody production peaks between 5 and 10 years of age, declining later in life.
• Elderly people may have undetectable anti-A and anti-B in reverse grouping.
• ABO antibodies cause rapid hemolysis if the incorrect ABO group is transfused and the patient may die.
• Anti-A (from group B) and anti-B (from group A) contain predominantly IgM, with small amounts of IgG.
• Group O individuals have anti-A, anti-B, and anti-A,B antibodies.
• Anti-A,B activity cannot be separated into pure specificities and is a separate "cross-reacting" antibody, usually IgG.
• Knowing IgG anti-A, anti-B, or anti-A,B amounts can predict or diagnose hemolytic disease of the fetus and newborn (HDFN).
• Reagent anti-A,B is not needed for routine ABO testing since monoclonal antisera have made anti-A and anti-B more sensitive.
• Anti-A,B is used for ABO confirmation of blood donors to verify group O units.

Inheritance of ABO Blood Groups

• Bernstein described that individuals inherit one ABO gene from each parent, determining RBC antigens.
• ABO genes follow Mendelian genetics and are codominant.
• A, B, or O gene occupies one position (locus) on each chromosome 9.
• The O gene is an amorph, producing no detectable antigen.
• Group O phenotype is an autosomal recessive trait with two nonfunctional O genes.
• Group A and B refer to phenotypes, while AA, BO, and OO denote genotypes.
• Phenotype and genotype are the same in an AB individual due to inheriting A and B genes.

Formation of A, B, and H Red Cell Antigens

• ABH antigens are formed by interaction of genes at three loci (ABO, Hh, and Se).
• These genes produce glycosyltransferases that add sugars to a basic precursor substance.
• A, B, and H antigens form from the same precursor with sugars attached by enzyme transferases.
• The H antigen is the precursor on which A and B antigens are made, found on chromosome 19.
• The H gene must be inherited to form ABO antigens on RBCs.
• The Se gene must form ABO antigens in secretions.
• The RBC precursor substance is type 2, with terminal galactose attached to N-acetylglucosamine in a beta 1 → 4 linkage.
• A type 1 precursor has a beta 1 → 3 linkage between galactose;
• ABH antigens develop early in fetal life but are weaker on newborns.

Interaction of Hh and ABO Genes

• Group O individuals inherit at least one H gene (HH or Hh) and two O genes.
• The H gene produces α-2-L-fucosyltransferase, which transfers L-fucose to the terminal galactose of type 2 chains.
• L-fucose is then the sugar responsible for H specificity.
• Blood group O has the highest concentration of H antigen.
• The H gene is in more than 99.99% of the population therefore the genotype hh is extremely rare.
• Bombay lacks ABH antigens due to inheriting hh genotype, that does not elicit a-2-L-fucosyltransferase production and H substance is not expressed on RBCs.
• The A gene (AA or AO) codes for a-3-N-acetylgalactosaminyltransferase, which transfers N-acetyl-D-galactosamine (GalNAc) sugar to the H substance, responsible for A specificity.
• The A gene elicits higher transferase concentrations, converting almost all H antigen to A antigen sites.
• Group B inherits B gene (BB or BO) that codes for a-3-D-galactosyltransferase, which attaches D-galactose (Gal) sugar to H substance.
• If both A and B genes are inherited, the B enzyme competes more efficiently for the H substance than the A enzyme.

Molecular Genetics of ABO

• More than 200 ABO alleles have been identified by molecular investigation.
• The ABO gene is located on chromosome 9 and consists of seven exons.
• Exons 6 and 7 are for the catalytic domain of ABO glycosyltransferases, most of the coding sequence in exon 7.
• Amino acid substitutions within these two exons are responsible for transferring the immunodominant sugar to H substance.
• All ABO antigens arise from mutations in the single ABO gene.
• Three specific mutations, which are more common, indirectly lead to changes in the epitope structures resulting in A, B, or O specificities.
• ABO genotyping is complex due to ABO locus diversity.

Formation of A, B, and H Soluble Antigens

• ABH antigens are in membranes of RBCs, endothelial cells, platelets, lymphocytes, and epithelial cells.
• Their presence is dependent on inherited ABO genes and Sese (secretor genes).
• Eighty percent are secretors (SeSe or Sese) they then regulate their formation.
• The Se gene produces a transferase (a-2-L-fucosyltransferase) that modifies the type 1 precursor substance in secretions, forming H substance and expressed as A and B substance.
• People who inherit the sese genotype are nonsecretors.
• Formation of soluble A, B, and H substances is the same as on RBCs, only with minor distinctions.
• ABH secretion tests were done to establish the true ABO group of an individual whose RBC antigens are poorly developed.
• The term secretor refers only to secretion of soluble A, B, and H antigens in bodily fluids.
• Types 1 and 3 are primarily associated with body secretions.
• Types 2 and 4 are associated with the red cell membrane.

ABO Subgroups

A Subgroups

• In 1911, differentiated two A antigens based on reactions between group A RBCs and anti-A and anti-A1.
• Group A RBCs reacting with both anti-A and anti-A1 classified as A1.
• Those reacting with anti-A but not anti-A1 classified as A2.

Concepts

• Weaker serologic reactivity of ABO subgroups is because of decreased A and B antigen sites on RBCs. Classification include into A1 and A2 phenotypes accounts for 99% of all group A individuals.
• The production of antigens depends on an inherited gene at the ABO locus.
• Qualitative differences also involves 1 to 8% of A2 individuals producing anti-A1 that can cause discrepancies in forward and reverses ABO testing.
• A2 is characterized by a single-base substitution at nucleotide 467 and a single-base substitution at nucleotide 1059.
• When identifying A1 and A2 phenotypes reagent from the seeds of the plant Dolichos biflorus serve as a source of anti-A1 and are called Lectins.

Lectins

• Seed extracts agglutinate human cells with some specificity.
• They agglutinate A1 (or A1B) cells but not agglutinating A2 (or A2B cells)
• They are used in blood banking.
• Lectin from extract the plant Ulex europaeus aligns the reactions of human anti-H anti-Cera.
• H antigen may not be detectable in group A1 individuals.

Weak B Subgroups

• Very rare and much less frequent than A subgroups.
• They recognized as variations in the strength reaction against anti-B and anti-A,B
• Inheritance is similar to those of A Subgroups
• To be result alterante alleles at the B locus.