Introduction
classification of blood based on inherited differences (polymorphisms) in antigens on the surfaces of the red blood cells (erythrocytes). Inherited differences of white blood cells (leukocytes), platelets (thrombocytes), and plasma proteins also constitute blood groups, but they are not included in this discussion.
Historical background
The human ABO blood groups were discovered by Austrian-born American biologist Karl Landsteiner in 1901. Landsteiner found that there are substances in the blood, antigens and antibodies, that induce clumping of red cells when red cells of one type are added to those of a second type. He recognized three groups—A, B, and O—based on their reactions to each other. A fourth group, AB, was identified a year later by another research team. Red cells of the A group clump with donor blood of the B group; those of the B group clump with blood of the A group; those of the AB group clump with those of the A or the B group because AB cells contain both A and B antigens; and those of the O group do not generally clump with any group, because they do not contain either A or B antigens. The application of knowledge of the ABO system in blood transfusion practice is of enormous importance, since mistakes can have fatal consequences.The discovery of the Rh system by Landsteiner and Alexander Wiener in 1940 was made because they tested human red cells with antisera developed in rabbits and guinea pigs by immunization of the animals with the red cells of the rhesus monkey Macaca mulatta.
The importance of antigens and antibodies
The red cells of an individual contain antigens on their surfaces that correspond to their blood group and antibodies in the serum that identify and combine with the antigen sites on the surfaces of red cells of another type. The reaction between red cells and corresponding antibodies usually results in clumping—agglutination—of the red cells; therefore, antigens on the surfaces of these red cells are often referred to as agglutinogens.
Antibodies are part of the circulating plasma proteins known as immunoglobulins, which are classified by molecular size and weight and by several other biochemical properties. Most blood group antibodies are found either on immunoglobulin G (IgG) or immunoglobulin M (IgM) molecules, but occasionally the immunoglobulin A (IgA) class may exhibit blood group specificity. Naturally occurring antibodies are the result of immunization by substances in nature that have structures similar to human blood groups. These antibodies are present in an individual despite the fact that there has been no previous exposure to the corresponding red cell antigens—for example, anti-A in the plasma of people of blood group B and anti-B in the plasma of people of blood group A. Immune antibodies are evoked by exposure to the corresponding red cell antigen. Immunization (i.e., the production of antibodies in response to antigen) against blood group antigens in humans can occur as a result of pregnancy, blood transfusion, or deliberate immunization. The combination of pregnancy and transfusion is a particularly potent stimulus. Individual blood group antigens vary in their antigenic potential; for example, some of the antigens belonging to the Rh and ABO systems are strongly immunogenic (i.e., capable of inducing antibody formation), whereas the antigens of the Kidd and Duffy blood group systems are much weaker immunogens.
The blood group antigens are not restricted solely to red cells or even to hematopoietic tissues. The antigens of the ABO system are widely distributed throughout the tissues and have been unequivocally identified on platelets and white cells (both lymphocytes and polymorphonuclear leukocytes) and in skin, the epithelial (lining) cells of the gastrointestinal tract, the kidney, the urinary tract, and the lining of the blood vessels. Evidence for the presence of the antigens of other blood group systems on cells other than red cells is less well substantiated. Among the red cell antigens, only those of the ABO system are regarded as tissue antigens and therefore need to be considered in organ transplantation.
Chemistry of the blood group substances
The exact chemical structure of some blood groups has been identified, as have the gene products (i.e., those molecules synthesized as a result of an inherited genetic code on a gene of a chromosome) that assist in synthesizing the antigens on the red cell surface that determine the blood type. Blood group antigens are present on glycolipid and glycoprotein molecules of the red cell membrane. The carbohydrate chains of the membrane glycolipids are oriented toward the external surface of the red cell membrane and carry antigens of the ABO, Hh, Ii, and P systems. Glycoproteins, which traverse the red cell membrane, have a polypeptide backbone to which carbohydrates are attached. An abundant glycoprotein, band 3, contains ABO, Hh, and Ii antigens. Another integral membrane glycoprotein, glycophorin A, contains large numbers of sialic acid molecules and MN blood group structures; another, glycophorin B, contains Ss and U antigens.
The genes responsible for inheritance of ABH and Lewis antigens are glycosyltransferases (a group of enzymes that catalyze the addition of specific sugar residues to the core precursor substance). For example, the H gene codes for the production of a specific glycosyltransferase that adds l-fucose to a core precursor substance, resulting in the H antigen; the Le gene codes for the production of a specific glycosyltransferase that adds l-fucose to the same core precursor substance, but in a different place, forming the Lewis antigen; the A gene adds N-acetyl-d-galactosamine (H must be present), forming the A antigen; and the B gene adds d-galactose (H must be present), forming the B antigen. The P system is analogous to the ABH and Lewis blood groups in the sense that the P antigens are built by the addition of sugars to precursor globoside and paragloboside glycolipids, and the genes responsible for these antigens must produce glycosyltransferase enzymes.
The genes that code for MNSs glycoproteins change two amino acids in the sequence of the glycoprotein to account for different antigen specificities. Additional analysis of red cell membrane glycoproteins has shown that in some cases the absence of blood group antigens is associated with an absence of minor membrane glycoproteins that are present normally in antigen-positive persons.
Identification of blood groups
The basic technique in identification of the antigens and antibodies of blood groups is the agglutination test. Agglutination of red cells results from antibody cross-linkages established when different specific combining sites of one antibody react with antigen on two different red cells. By mixing red cells (antigen) and serum (antibody), either the type of antigen or the type of antibody can be determined depending on whether a cell of known antigen composition or a serum with known antibody specificity is used.
In its simplest form, a volume of serum containing antibody is added to a thin suspension (2–5 percent) of red cells suspended in physiological saline solution in a small tube with a narrow diameter. After incubation at the appropriate temperature, the red cells will have settled to the bottom of the tube. These sedimented red cells are examined macroscopically (with the naked eye) for agglutination, or they may be spread on a slide and viewed through a low-power microscope.
An antibody that agglutinates red cells when they are suspended in saline solution is called a complete antibody. With powerful complete antibodies, such as anti-A and anti-B, agglutination reactions visible to the naked eye take place when a drop of antibody is placed on a slide together with a drop containing red cells in suspension. After stirring, the slide is rocked, and agglutination is visible in a few minutes. It is always necessary in blood grouping to include a positive and a negative control for each test.
An antibody that does not clump red cells when they are suspended in saline solution is called incomplete. Such antibodies block the antigenic sites of the red cells so that subsequent addition of complete antibody of the same antigenic specificity does not result in agglutination. Incomplete antibodies will agglutinate red cells carrying the appropriate antigen, however, when the cells are suspended in media containing protein. Serum albumin from the blood of cattle is a substance that is frequently used for this purpose. Red cells may also be rendered specifically agglutinable by incomplete antibodies after treatment with such protease enzymes as trypsin, papain, ficin, or bromelain.
After such infections as pneumonia, red cells may become agglutinable by almost all normal sera because of exposure of a hidden antigenic site (T) as a result of the action of bacterial enzymes. When the patient recovers, the blood also returns to normal with respect to agglutination. It is unusual for the red cells to reflect antigenicity other than that determined by the individual's genetic makeup. The presence of an acquired B antigen on the red cells has been described occasionally in diseases of the colon, thus allowing the red cell to express an antigenicity other than that genetically determined. Other diseases may alter immunoglobulins; for example, some may induce the production of antibodies directed against the person's own blood groups (autoimmune hemolytic anemia) and thus may interfere with blood grouping. In other diseases a defect in antibody synthesis may cause the absence of anti-A and anti-B antibody.
Uses of blood grouping
1.Transfusion:The blood donated by healthy persons is tested to ensure that the level of hemoglobin is satisfactory and that there is no risk of transmitting certain diseases, such as AIDS or hepatitis. It is then fractionated (split) into its component parts, particularly red cells, plasma, and platelets. Correct matching for the ABO system is vital. Compatible donors on the basis of their possessing A, B, or O blood are shown in the table.
As explained above, the most important blood group systems for transfusion of red cells are ABO and Rh. Persons who have either of the red cell antigens (A and B) have antibody present in their serum of the type that will oppose an antigen of its opposite nature; for example, group A blood contains A antigens on red cell surfaces and anti-B antibodies in the surrounding serum. On the other hand, O group individuals lack both the A and the B antigen and thus have both anti-A and anti-B in their serum. If these antibodies combine with the appropriate antigen, the result is hemolytic transfusion reaction and possibly death. Red cell transfusions must therefore be ABO compatible. The blood groups A and B have various subgroups (e.g., A1, A2, A3, and B1, B2, and B3). The only common subgroups that are likely to affect red cell transfusions are the subgroups of A.
Potential donors are also tested for some of the antigens of the Rh system, since it is essential to know whether they are Rh-positive or Rh-negative. Rh-negative indicates the absence of the D antigen. Rh-negative persons transfused with Rh-positive blood will make anti-D antibodies from 50 to 75 percent of the time. Antibody made in response to a foreign red cell antigen is usually not harmful but does require subsequent transfusions to be antigen-negative. Rh-positive blood should never be given to Rh-negative females before or during the childbearing age unless Rh negative blood is not available and the transfusion is lifesaving. If such a woman subsequently became pregnant with an Rh-positive fetus, she might form anti-Rh antibody, even though the pregnancy was the first, and the child might develop erythroblastosis fetalis (hemolytic disease of the newborn).
Care must be taken not to give a transfusion unless the cells of the donor have been tested against the recipient's serum. If this compatibility test indicates the presence of antibodies in the recipient's serum for the antigens carried by the donor's cells, the blood is not suitable for transfusion because an unfavourable reaction might occur. The test for compatibility is called the direct match test. It involves testing the recipient's serum with the donor's cells and by the indirect Coombs test. These are adequate screening tests for most naturally occurring and immune antibodies.
If, in spite of all the compatibility tests, a reaction does occur after the transfusion is given (the unfavourable reaction often manifests itself in the form of a fever), an even more careful search must be made for any red cell antibody that might be the cause. A reaction after transfusion is not necessarily due to red cell antigen-antibody reactions. It could be caused by the presence of antibodies to the donor's platelets or white cells. Transfusion reactions are a particular hazard for persons requiring multiple transfusions.
2.Organ transplants:The ABO antigens are widely distributed throughout the tissues of the body. Therefore, when organs such as kidneys are transplanted, most surgeons prefer to use organs that are matched to the recipient's with respect to the ABO antigen system, although the occasional survival of a grafted ABO-incompatible kidney has occurred. The remaining red cell antigen systems are not relevant in organ transplantation.
Blood groups and disease
In some cases an increased incidence of a particular antigen seems to be associated with a certain disease. Stomach cancer is more common in people of group A than in those of groups O and B. Duodenal ulceration is more common in nonsecretors of ABH substances than in secretors. For practical purposes, however, these statistical correlations are unimportant. There are other examples that illustrate the importance of blood groups to the normal functions of red cells.
In persons who lack all Rh antigens, red cells of altered shape (stomatocytes) and a mild compensated hemolytic anemia are present. The McLeod phenotype (weak Kell antigens and no Kx antigen) is associated with acanthocytosis (a condition in which red cells have thorny projections) and a compensated hemolytic anemia. There is evidence that Duffy-negative human red cells are resistant to infection by Plasmodium knowlesi, a simian malaria parasite. Other studies indicate that P. falciparum receptors may reside on glycophorin A and may be related to the Wrb antigen.
Blood group incompatibility between mother and child can cause erythroblastosis fetalis (hemolytic disease of the newborn). In this disease IgG blood group antibody molecules cross the placenta, enter the fetal circulation, react with the fetal red cells, and destroy them. Only certain blood group systems cause erythroblastosis fetalis, and the severity of the disease in the fetus varies greatly. ABO incompatibility usually leads to mild disease. Rh, or D antigen, incompatibility is now largely preventable by treating Rh-negative mothers with Rh immunoglobulin, which prevents immunization (forming antibodies) to the D antigen. Many other Rh antigens, as well as other red cell group antigens, cause erythroblastosis fetalis. The baby may be anemic at birth, which can be treated by transfusion with antigen-negative red cells. Even total exchange transfusion may be necessary. In some cases, transfusions may be given while the fetus is still within the uterus (intrauterine transfusion). Hyperbilirubinemia (an increased amount of bilirubin, a breakdown product of hemoglobin, in the blood) may lead to neurological deficits. Exchange transfusion eliminates most of the hemolysis by providing red cells, which do not react with the antibody. It also decreases the amount of antibody and allows the child to recover from the disease. Once the antibody disappears, the child's own red cells survive normally.