Blood Group Systems

International Society of Blood Transfusion (ISBT) Working Party has organized red cell antigens into 25 blood group systems. Red cell antigens that are produced by alleles (alternative forms of a specified gene) at a single gene locus or very closely linked loci constitute a blood group system.

Blood group genes are inherited in a Mendelian manner and are mostly located on autosomes. Most of the blood group genes are expressed in a co-dominant manner (i.e. the two allelic forms are expressed equally if inherited in a heterozygous state, and no gene or allele is dominant over another). The particular alleles at a specified gene locus in an individual constitute the genotype. The phenotype is the outward expression of the genotype.

In blood transfusion practice, most important blood group systems are ABO and Rh. This is because, A, B, and Rh D antigens are the most immunogenic (i.e. capable of eliciting a strong antibody response on stimulation) and their alloantibodies can cause the destruction of transfused red cells or induce hemolytic disease of the newborn (HDN). ABO antigens are also important in organ transplantation.

ABO SYSTEM

In ABO system, there are four main types of blood groups: A, B, AB, and O. Identification of these four blood groups is based on presence or absence of A and/or B antigens on red cells. According to Landsteiner’s law, anti-A and/or anti-B antibodies are always present in plasma of individuals who lack corresponding antigen(s) on their red cells (Table 1200.2). ABO is the only blood group system, in which if an antigen is absent in an individual, the corresponding antibody is always present in plasma.

There is a geographical variation in frequencies of different ABO blood groups. However, generally, the O blood group is the most common, while the AB group is the least common in a population. Read also: ABO Grouping and Rh D Grouping Methods

Table 1200.1: Blood group systems

ABO	Dombrock
MNS	Colton
P	Landseiner-Weiner
Rh	Chido/Rogers
Lutheran	Hh
Kell	Kx
Lewis	Gerbich
Duffy	Cromer
Kidd	Knops
Diego	Indian
Yt	Ok
Xg	Raph
Scianna	-

Note: A and B genes are dominant while the O gene is recessive. Individuals with AA, BB, and OO are homozygous respectively for A, B, and O, while AO, BO, and AB are heterozygous.

Blood Group O

This is usually the most frequently occurring blood group in the population. Group O red cells have large amounts of H antigen on their surface. Genotype is OO and the antibodies in plasma are anti-A, anti-B, and anti-A,B. These antibodies are usually IgG in nature. They can, therefore, cross the placenta and induce hemolytic disease of the newborn.

Blood Group A

Genotype of group A persons is either AA or AO, and the antigens present on the red cells are A and H. Their plasma contains anti-B antibodies that are IgM in nature. Blood group A is divided into two subgroups: A1 (80%) and A2 (20%). Anti-A1 produced by A2 or A2B individuals is weak and clinically insignificant; however, it can cause problems during blood grouping. Some persons with A2 and with A2B can make anti-A1. A1 and A2 red cells can be differentiated by lectins of Dolichos biflorus that cause agglutination of A1 red cells, but not of A2 red cells.

Blood Group B

Genotype of group B persons is either BB or BO, and the antigens present on red cells are B and H. Their plasma contains anti-A antibodies that are IgM in nature.

Blood Group AB

This is the least common of the ABO blood groups. Genotype is AB, and antigens present on red cells are A, B, and very small amount of H. The AB blood group is further subdivided into A1B and A2B. Plasma of group AB individuals contain no anti-A or anti-B antibodies.

Antigens of the ABO system

Antigens of the ABO system are: A (A1, A2), B, and H. They are glycolipids, glycoproteins, or glycosphingolipids. In addition to red cells, they are also expressed on white cells, platelets, and body tissues. They are also present in a soluble form in various body secretions (in secretors) (Table 1200.3).

Table 1200.3: Distribution of ABH antigens

Red blood cells White blood cells (weak) Platelets (weak) Body tissues Body fluids (in secretors)

ABO antigens are poorly expressed at birth, increase gradually in strength and become fully expressed around 1 year of age. In older age, they become slightly weak.

Formation of ABH Antigens

The H gene (genotype HH or Hh) produces a transferase enzyme, which changes precursor substance (present on red cells) into H substance. The A and B genes (chromosome 9) produce specific transferase enzymes, which convert H substance into A and B antigens respectively.

Sugar N-acetylgalactosamine is added to H antigen chain to make the A antigen, while sugar galactose is added to H antigen chain to produce the B antigen. Some amount of H antigen remains unchanged on red cells. The O gene produces an inactive transferase so that H antigen persists unchanged on red cells (Figure 1200.1).

Some persons do not inherit the H gene (genotype hh) and thus cannot synthesize H substance. Such persons may inherit the A or B gene but cannot express it, as they are unable to produce the H substance. Such individuals are said to have Bombay phenotype or Bombay blood group (Oh) (the genotype of Bombay phenotype is hh). The name derives its name because it was first detected in persons living in Bombay, India. It has also been described in other populations. This blood group is extremely rare. There is complete lack of H, A, and B antigens due to the lack of H (and Se) genes. Their red cells type as group O; however, unlike group O individuals, Oh persons have no H antigen on their red cells and their plasma contains strong anti-H in addition to anti-A and anti-B. On cross-matching, all units are incompatible (Table 1200.4). Therefore Bombay group persons should be transfused only with Oh blood.

Table 1200.4: Identification of Bombay phenotype

Blood group: O Cross-matching: Serum incompatible with O cells Red cells yield negative reaction with anti-H lectin

Secretors and Non-secretors

Secretors are persons who secrete A, B, and H antigens into body fluids (such as plasma, gastric juice, saliva, sweat, tears, semen, milk, etc.). This ability is dependent on presence of a dominant secretor gene (Se) on chromosome 19. About 80% of individuals are secretors (genotype Sese or SeSe) and remaining are non-secretors (genotype sese). Both secretors and non-secretors express ABO antigens on red cells.

Antigens secreted by different ABO blood groups are:

• Group A: A, H
• Group B: B, H
• Group AB: A, B, H
• Group O: H

Antigens secreted into body fluids are called as ABH substances. Testing for ABH substances in saliva may be helpful when red cell grouping yields uncertain results.

Determining secretor status in saliva and semen can be helpful in resolving ABO blood group discrepancies and in forensic studies (e.g. semen sample collected from a rape victim revealing a soluble ABH antigen that does not match with the ABO blood group of the accused). Inhibitor tests are used to detect the presence of soluble blood group antigens in body secretions. If saliva contains a soluble antigen, and if corresponding antibody is added, the activity of the antibody is neutralized. When red cells carrying the appropriate antigens are subsequently added to the mixture, there will be inhibition of agglutination (i.e. the person is a secretor). If agglutination occurs, then the individual is a non-secretor.

Example:

1. Blood group: B
   Saliva + anti-B; Add B cells: No agglutination
   Interpretation: Secretor
2. Blood group: O
   Saliva + Anti-A; Add A cells: Agglutination; Saliva + Anti-B; Add B cells: Agglutination; Saliva + Anti-H;
   Add O cells: No agglutination.
   Interpretation: Secretor
3. In non-secretors, no soluble antigens are present in secretions. Thus antibodies in the reagent are free to bind to red cells of respective group. There will be agglutination reaction.

Antibodies of the ABO System

The most important antibodies in transfusion practice are anti-A and anti-B. They are also called as naturally occurring antibodies because they arise without immune stimulation by red cells by relevant blood group antigens.

They are regularly-occurring, i.e. if an antigen is absent the corresponding antibody is always present. They are not detectable in the blood of newborn infants due to their underdeveloped immune system and appear around 3-6 months of life. It is thought that they are produced in response to A- and B-like antigens of bacteria, which are present in the intestine and certain foods. If anti-A and/or anti-B are present at birth, they are of maternal origin (IgG). Anti-A and anti-B antibodies are usually of IgM class. Immune or IgG antibodies (anti-A or anti-B) can be stimulated by exposure to red cells, white cells, or platelets.

IgM antibodies are large molecules and can bind up to ten antigens; therefore they can cause direct agglutination of red cells and cause visible agglutination. IgM antibodies can efficiently fix the complement. Naturally occurring ABO antibodies can cause:

• Hemolytic transfusion reaction in a case of ABOmismatched blood transfusion,
• Acute graft rejection in case of ABO-incompatible solid organ transplantation, and
• Hemolysis of donor red cells following ABOincompatible bone marrow transplantation.

Less commonly, some individuals have large amounts of ABO antibodies of immune nature. Usually, group O individuals following immune stimulation by transfusion, pregnancy, or injection of certain vaccines or toxoids (that contain bacterial A- and B-like antigens) produce them in large amounts. These antibodies are of IgG class, of high titer, and cannot be neutralized by soluble blood group antigens. If blood of such group O individuals (called dangerous universal group O donors) is transfused to group A or B individuals, serious hemolysis of recipient’s red cells can occur. Therefore group O donors should not be employed as universal donors. In addition to causing hemolytic transfusion reaction, these IgG antibodies can cross the placenta and induce hemolytic disease of newborn.

Concepts of Universal Donor and Recipient

Red cells of group O donors are devoid of A and B antigens and cannot be agglutinated by anti-A and anti-B antibodies. Therefore, group O persons are traditionally considered as universal donors.Group AB persons are considered as universal recipients. This, however, is an oversimplification because only the reaction between recipient’s plasma (antibodies) and the donor’s red cells is considered; a small amount of antibodies in donor’s plasma can cause destruction of recipient’s red cells. In addition, antigens other than A, B, or RhD can bind to corresponding antibodies or immunize the recipient.

THE Rh SYSTEM

When Rhesus monkey red cells were injected into rabbits and guinea pigs (by Landsteiner and Weiner in 1940), antibody, which was raised, was found to react with Rhesus monkey red cells as well as with 85% of human red cells (White residents of New York city). The antigen involved was called as Rh factor. Subsequently it was shown that the original antibody was different from anti-D antibody discovered later. The name of the antigen, however, has remained as Rhesus. Apart from Landsteiner and Weiner, credit for discovery of Rh system also goes to Levine and Stetson who discovered in 1939 the antibody (actually anti-D) that caused hemolytic disease of newborn.

The Rh system is only next in importance to ABO system in transfusion practice. The importance of this system lies in the high immunogenicity of Rh D antigen, which readily induces formation of anti-D antibodies in 50-70% of Rh D-negative individuals. Anti-D antibodies can cause hemolytic transfusion reaction or, in pregnant women, Rh hemolytic disease of newborn.

The Rh system was discovered independently by: (1) Stetson and Levine in 1939 and (2) by Landsteiner and Weiner in 1940.

Antigens of the Rh System

Rh system is highly complex and consists of about 40 antigens. The important antigens of the Rh system are C, D, E, c, and e. Antigen d does not exist. D antigen is the most immunogenic. There are various nomenclature systems for Rh antigens. Fisher-Race or CDE nomenclature system and Weiner system are popular.

According to Fisher and Race, three closely linked genes are inherited together on one chromosome (haplotype) from each parent. Allelic forms of these genes are C and c, D and d, and E and e with eight possible haplotypes: Cde, cde, cDE, cDe, cdE, Cde, CDE, and CdE. As an individual inherits one haplotype from each parent, 36 genotypes are possible such as Cde/cde, Cde/cDe, CDE/cde, etc (Figure 1200.2). The presence of D in either homozygous (D/D) or heterozygous (D/d) state makes that individual Rh positive, while Rh negative persons are homozygous for d (d/d). It was thought that d gene was an amorph.

According to the theory by Dr. Alexander Weiner, a single Rh gene is inherited from each parent; this single Rh gene, however, has multiple alleles (Figure 1200.3). The Weiner system uses Rh-Hr nomenclature. The major difference between Fisher-Race and Weiner systems is that according to Fisher-Race, there are three closely linked genes that are inherited from each parent, while according to Weiner, a single gene with multiple alleles is inherited from each parent.

Results of current genetic studies suggest that both Fisher-Race and Weiner systems are partially correct. It has been found that the RH locus is located on chromosome 1 and consists of two closely linked genes-RHD and RHCE (Figure 1200.4). The alleles of RHCE are CE, Ce, ce, and cE.

In Rh negative persons, deletions, point mutations, or partial mutations of D gene have been found. Rh antigens are expressed only on red cells and not on any other tissues. They are also not secreted in body fluids. In contrast to ABO antigens, Rh antigens are fully expressed on red cells before birth and also on red cells of early fetuses.

Depending on the presence or absence of antigen D on red cells, a person is grouped either as Rh positive (when red cells express antigen D) or Rh negative (when D antigen is absent on red cells). Frequency of D antigen varies in different populations. In India, approx. 95% of the people express D antigen on their red cells (Rh D positive), while 5% are Rh D negative. The frequency in Caucasians is 85% Rh positive and 15% Rh negative.

Other forms of D antigen are weak D and partial D (Figure 1200.5). Red cells having weak D antigen were formerly called as D^u cells which react weakly with anti-D reagent. There is a quantitative reduction in the number of D antigen sites on such red cells. D^u recipients do not make anti-D antibodies following stimulation by D antigen (e.g. following D positive blood transfusion). D^u donors should be considered as Rh positive and their blood should not be transfused to Rh negative donors.

In red cells having partial D antigen, parts of D antigen are missing. Variants of partial D antigen exist. Individuals with D^VI variant are able to produce anti-D antibody against the missing part of the antigen. Such recipients should be considered as Rh negative, while donors should be regarded as Rh positive. However, in practice, individuals with partial D antigen are typed as D negative and are identified only after they have produced anti-D antibodies.

Complete absence of all Rh antigens on red cells (Rh null cells) is associated with stomatocytosis and compensated hemolysis.

Rh Antibodies

In general, most Rh antibodies are of immune type, i.e. they are the result of immunization by blood transfusion or pregnancy. Most of these antibodies are of IgG class.

Rh antibodies can cause hemolytic transfusion reaction or hemolytic disease of newborn (HDN). Since Rh antibodies do not activate complement, hemolysis is extravascular and predominantly occurs in spleen. Due to the high immunogenicity of D antigen, Rh negative persons (especially women of childbearing age) should be transfused only with Rh-negative blood. During pregnancy, IgG anti-D can cross the placenta and induce hemolytic disease of newborn by causing immune hemolysis of fetal red cells. Rh hemolytic disease of newborn can be prevented by prophylactic administration of Rh immune globulin to all Rh-negative women during mid pregnancy and within 72 hours of delivery. Anti-D and anti-c can cause severe HDN. Anti-C, anti-E, and anti-e usually do not cause HDN or cause mild HDN. Relative immunogenicity of Rh antigens is shown in Figure 1200.6.

OTHER BLOOD GROUP SYSTEMS

Salient features of some other blood group systems are summarized in below table.

Table 1200.5: Blood group systems other than ABO and Rh

Blood Group System	Antigens	Antibodies	Comments
1. Lewis	Lea, Leb	Natural, IgM	Lewis antigens are passively absorbed from plasma on red cells; Lewis antibodies are rarely of clinical significance
2. Kell	About 20 (KEL1, KEL2, etc)	IgG	Anti-K antibodies can cause hemolytic transfusion reaction and hemolytic disease of newborn; Some individuals do not have precursor substance on their red cells from which K antigens are produced; such red cells have short life, acanthocytic features, and express Kell antigens weakly (MacLeod phenotype).
3. Duffy	Fya, Fyb	IgG	Antibodies can cause hemolytic transfusion reaction. Plasmodium vivax enters the red cells at the Duffy antigen site. The Fy(a-b-) phenotype in blacks confers resistance against Plasmodium vivax infection
4. Kidd	JKa, JKb	IgG	Antibodies cause delayed transfusion reaction and mild hemolytic disease of newborn
5. MNSs	M, N, S, s	-	M and N antigens are important in paternity testing
6. P	P, P1, Pk	IgM, IgG	Auto-anti-P occurs in paroxysmal cold hemoglobinuria