Chapter IX - Medical Biology - Genetics 2023-2024 PDF

Exercise 9 Topic: Human blood group systems. Glossary: Agglutination - clumping reaction: the formation of a solid mass by aggregating suspended particles in a solution. An antigen (agglutinogen) is bound by agglutinins (antibodies, lectins), which results in the formation of large, precipitating co...

Exercise 9 Topic: Human blood group systems. Glossary: Agglutination - clumping reaction: the formation of a solid mass by aggregating suspended particles in a solution. An antigen (agglutinogen) is bound by agglutinins (antibodies, lectins), which results in the formation of large, precipitating complexes; agglutination tests are used in the diagnosis of blood group determination and determining the possibility of transfusion (red blood cell agglutination). Antigen - a molecule or a multi-component substance that is recognized by the competent cells of the immune system, which can result in the activation of the immune response in the production of specific antibodies and / or the creation of a specific cellular response; a molecule that reacts specifically with an antibody or a sensitized cell; a complex antigenic molecule may carry several antigenically distinct sites (determinants). Antibody (immunoglobulin) - proteins (glycoproteins), mainly from the gamma-globulin group, secreted in the course of the humoral immune response; they have the ability to bind antigens specifically; the immunological specificity of the antibody is conditioned by the specific amino acid sequence and thus the spatial arrangement of variable fragments (the antigen binding site) of the antibody-building polypeptide chains. As a result of the reaction of antibodies with antigens we observe different reactions, e.g. agglutination, or lysis of antigen-bearing cells. Endogenous antigens (natural antigens) - are generated within normal body cells and then presented on the cell surface (directly – erythrocyte antigens) or with the help of major histocompatibility complex (MHC) proteins which cause that the immune system of each body recognizes and accepts its own antigens, and destroys foreign (distinguishes between its own and foreign substances, cells). Immune alloantibodies (irregular) - produced by the human body as a result of immune response in case of contact with antigenically foreign blood cells, which were in the circulation (e.g after incompatible blood transfusion or, as a result of a serological conflict in the Rh system during pregnancy: immunologic antibodies Rh system belonging to the IgG class). Lectins - proteins or glycoprotein-binding glycoproteins widely distributed in seeds and other parts of plants, as well as in almost all organisms (bacteria to mammals); they bind to sugar-specific residues of glycoproteins present on the surface of cells (e.g. erythrocytes, lymphocytes) causing their clumping (agglutination, phytoagglutination). Natural (regular) antibodies - occurring in every human being, appearing naturally in the first year after birth; present in the blood throughout life: antibodies to ABO system antigens - natural isohemagglutinins of ABO system against antigen A (anti-A) and against antigen B (anti-B); they belong to the IgM class. Blood groups The first attempts to treat blood transfusions began in the 17th century, when the English physician William Harvey first described in scientific terms the blood circulation in the human body (1667 Jean Baptiste Denis - court physician to Louis XIV, transfused about 270 ml of arterial lamb’s blood into the veins of the boy’s arm, after earlier blood draining to heal his high fever, the boy survived the procedure, and even recovered). For many consecutive years, especially in the case of massive haemorrhages, blood transfusions were carried out, ending with both positive results and failures. Discovery of the ABO group system (Karl Landsteiner, 1901) and description of agglutination (clumping) of red blood cells in contact with antibodies present in blood plasma of groups with different antigenic structure, as well as overcoming difficulties related to blood coagulation (1914), due to the use of sodium citrate (which is the basic ingredient of most solutions preserving blood and its components to this day), has begun a new chapter in the field of transfusiology. As a result of incompatible blood transfusion, intravascular destruction of red blood cells occurs, often with complement. Early recognition of blood groups determines the possibility of preventing such a phenomenon. There are numerous polymorphic genes, glycoproteins and glycolipids on the surface of human red blood cells. They are blood group antigens which are inherited according to Mendel's laws and are usually detected by serological tests. The agglutination technique made it possible to detect 36 major human blood group systems and at least 322 distinct red blood cell antigens, also 38 antigens not related to known group systems. Among the group of antigens we distinguish antigens: private - with a low 99% incidence. The use of serological and molecular methods still allows to discover new clinically relevant blood group antigens. In the inheritance of the antigens of erythrocyte group systems, there are phenomena of gene cooperation, autosomal linkage, or linkage with the X chromosome (as well as gene polymorphism). AB(O)H system The AB(O)H system antigens found on the membranes of erythrocytes and other cells of the human body (except hepatocytes and nerve tissue cells) are conditioned by the presence of three major alleles: IA, IB, i (I0) with the locus on the 9th pair of chromosome and the H, h alleles, with the locus on chromosome 19. Antigens A, B, H can be also found in body fluids and human body secretions (e.g. saliva, milk, sweat, amniotic fluid, urine, faeces). It is dependent on the dominant Se allele with the locus on 19th chromosome pairs (linked to the H allele). The AB(O)H system antigens are carbohydrates - they are not direct gene products. The products of all of the above-mentioned alleles are enzymes - glycosyltransferases carrying the respective sugar, attached to the end of a particular type of oligosaccharide chain thus forming an antigen: H, when enzyme α-2-L-fucosyltransferase carries L-fucose; A, when enzyme α-3-N-acetyl-D-galactosaminotransferase carries N-acetylgalactosamine; B, when enzyme α-3-D-galactosyltransferase carries galactose. H antigen is necessary for the formation of A and B antigens. On the cell membranes of erythrocytes it is formed by the action of α-2-L-fucosyltransferase FUT1 encoded by the gene H. In contrast, in the secretory epithelial cells, a soluble form of H antigen is formed. This phenomenon is dependent on the Se gene responsible for the production of α2-L-fucosyltransferase FUT2, differing in amino acid sequence from FUT1 (both genes show 70% nucleotide homology; sequence differences most likely are caused by duplication). The FUT2 enzyme also determines the synthesis of Lewis blood system antigens. The absence of H antigen occurs in about 0.0004% (4: 1,000,000) of the human population (in India, 0.01% of the population). For the first time, such a case was described in 1952 in Bombay, now known as Mumbai, in a woman in need of a blood transfusion. The absence of A and B antigens on erythrocytes indicated O blood group. However, one of the patient's parents had AB, and two of her children - B group. It was found that the woman was homozygous for recessive mutation of the H gene. She had the genotype IBi hh, which was defined as the Bombay Oh phenotype. Currently, we distinguish the classic Bombay and para-Bombay phenomena (Table 1). Table 1. Comparison of the classic Bombay and para-Bombay phenotype. The classic Bombay phenotype No expression of H antigen on erythrocytes No H antigen in saliva (in body fluids and secretions of the human body) The presence of anti-H, anti-A, anti-B antibodies in plasma Genotype: hh sese The para-Bombay phenotype No expression of H antigen on erythrocytes The presence of H antigen in saliva (depending on the genotype, the presence of H, A and B antigens in saliva, as well as in plasma and their adsorption on erythrocytes) The presence of anti-H antibodies in plasma Genotype: hh Sese / hh SeSe Example: Proband (mother), whose parents had blood groups A and AB, gave birth to three children with groups: A, B, AB. Soon, she needed a blood transfusion due to a serious injury. In a blood test the absence of A and B antigens on erythrocytes indicated O blood group. Table 2. Genotypes Phenotypes Phenotypes - offspring Genotypes - offspring Mother ♀ IAIB Hh Sese Mother ♀ AB ̅̅̅ 𝑆𝑒 AB ̅̅̅ 𝑆𝑒 IAIB Hh Sese IAIB Hh SeSe IAIB HH Sese IAIB HH SeSe A ̅̅̅ 𝑆𝑒 IA i Hh Sese IAIA Hh SeSe IAIA HH Sese IAIA HH SeSe x x B ̅̅̅ 𝑆𝑒 IB i Hh Sese IB i Hh SeSe IB i HH Sese IB i HH SeSe IAi Hh Sese ♂ Father ̅̅̅ A 𝑆𝑒 ♂ Father O ̅̅̅ 𝑆𝑒 IA i hh Sese IA i hh SeSe IB i hh Sese IB I hh SeSe IBIB hh SeSe IBIB hh Sese IAIB hh SeSe IAIB hh Sese IAIA hh Sese IAIA hh SeSe Oh 𝑠𝑒 ̅̅̅ IA i hh sese IB i hh sese IAIA hh sese IAIB hh sese The gene polymorphism of the ABO system determines the occurrence of antigen varieties (designated A1, A2, A3, Ax, Am, Ael, Aend, etc.), which have a correspondingly weaker expression of A antigen. Similarly, although less often, weak variants are detected in people with B group (e.g., B3, Bx, Bm, Bw). Alleles of these subgroups show single nucleotide polymorphisms (SNPs) in exons or promoter regions or contain mutations that cause amino acid substitutions or frame offsets. This results in a decrease in enzyme glycosyltransferases activity and a reduced number of antigen molecules on erythrocytes. Example: The A2 allele may differ in the coding region from the A1 allele in two ways: 1. substitution of a single nucleotide (467C> T) that results in a single amino acid change (proline at position 156 changed to leucine); 2. deletion (1061delC), which results in a shift of the reading frame comprising a fragment of 64 nucleotides (Yamamoto et al. 1992). The ABO system is the only in which there are natural blood serum antibodies against antigens, absent on the own erythrocytes. Antibodies ABO: anti-A and anti-B are produced in infancy (no earlier than 4 months of age) and are called natural (regular) antibodies. They belong to the IgM class; do not pass through the endothelium and the placenta. They are produced against antigens of bacteria naturally found in the human intestine (microbiome). The frequency of ABO system alleles and blood groups is different in individual human populations (Table 3, Figure 1). Table 3. Frequency of the ABO system allele in selected populations: Population The allele frequency (%) O (i) A (IA) B (IB) Americans 67 26 7 French 64 30 6 Japanese 55 28 17 Africans 57 22 21 Indians 55 18 26 Rh system The Rh system antigens, which are polypeptides, are found only on the membranes of erythrocytes. They depend on two genes: RHD and RHCE placed on the short arm of chromosome 1. The Rh system is the most polymorphic antigen system of red blood cells. Forty-nine antigens of this system were detected, however, 5 antigens are clinically relevant: D, C, c, E, e. The RHD gene encodes the D antigen and its variants, whereas RHCE - the other antigens. Determinants of antigen D are a component of one polypeptide, and determinants C, c, E and e are located in a second polypeptide chain. Differences between antigens C and c and E and e result from point mutations that cause changes in individual amino acids. The D-antigen is the strongest immunogen, from all known erythrocyte antigen systems. The presence of D-antigen determines the Rh+ phenotype. The absence of the RHD gene and also the D antigen is recorded as the dd genotype and the Rh- phenotype. This applies to the Caucasians (about 15% of the population), where the lack of the RHD gene most likely results from the deletion. In contrast, in Japanese people and representatives of the human black variety, the RHD gene is present but unable to produce protein, due to disturbed regulation of its activity. The use of diagnostic reagents (monoclonal reagents) with specificities of anti-D, -C, -c, -E, -e allowed to isolate 18 Rh phenotypes. Because all RH genes lie on one chromosome (linked genes), we say that the phenotype depends on two haplotypes from the parents (Table 4). Table 4. Examples of Rh phenotypes and possible genotypes: dccee Prevalence of phenotype in Poland [%] 16.17 DCcee 33.58 DCe/dce, DCe/Dce, Dce/dCe DCCee 15.22 DCe/DCe, DCe/dCe DccEe 12.82 DcE/dce, DcE/Dce, Dce/dcE DCcEe 10.03 DCE/Dce, DCE/dce, Dce/dCE, DCe/DcE, DCe/dcE, Dccee 3.51 Dce/dce, Dce/Dce Phenotype Possible genotypes (the certain or most probable genotype is shown) dce/dce The occurrence of blood groups in AB0 and Rh systems of selected populations in the world is presented in Table 5. Table 5. Frequency of blood groups of the AB0 and Rh systems in selected populations. Population Frequency of blood groups [%] 0+ A+ B+ AB+ 0- A- B- AB- Poland 31 32 15 7 6 6 2 1 USA 38 34 9 3 7 6 2 1 United Kingdom 37 35 8 3 7 7 2 1 Australia 40 31 8 2 9 7 2 1 Finland 27 38 15 7 4 6 2 1 Sweden 32 37 10 5 6 7 2 1 France 36 37 9 3 6 7 1 1 Fetomaternal antigen incompatibility RhD haemolytic disease of the newborn (also known as rhesus incompatibility, rhesus D haemolytic disease of the newborn, RhD HDN) The fetomaternal antigen incompatibility is the phenomenon of pregnant women producing antibodies directed against fetal antigens that the child inherited from their father, and the mother does not have them. Antigens occur on the surface of various blood cells, such as erythrocytes, platelets or granulocytes. The maternal immune system destroys fetal cells, which can even lead to death. Most often the incompatibility occurs in the Rh system. Women who do not produce D antigen (Rh negative blood group) may undergo alloimmunization induced by the RhD antigen present in the fetus. This process usually occurs during labour (the process of childbirth, especially the period from the start of uterine contractions to delivery), when fetal erythrocytes infiltrate the mother's bloodstream. For the initiation of the immune response in the mother, a leak of 0.1 ml of fetal blood suffices. The Rh incompatibility and haemolytic disease of the fetus / newborn usually relate to a second pregnancy in which the child also inherits the RhD antigen from the father. The mother produces anti-RhD IgG antibodies that go through the placenta and bind to the erythrocyte RhD antigen in the fetal bloodstream. Erythrocytes covered with antibodies - "coated" - are destroyed by cells of the phagocytic system (former name - reticuloendothelial system), which leads to anaemia, and in extreme cases to death of the fetus or newborn. In pregnancy, the level of anti-D antibodies in the mother's serum is determined several times: for the first time around the 12th week of pregnancy, then every six-eight weeks and always if there has been any bleeding or injury. The level of anti-D antibodies is also tested just after delivery. To prevent this incompatibility, the immunoprophylaxis was developed in 1966, in which mothers with the Rh (-) group in whom anti-RhD antibodies have not been detected up to 72 hours after the delivery of a child with Rh (+), are given IgG anti-D immunoglobins (destruction of blood cells with antigen D). The incidence of the feto-maternal antigen D incompatibility has thus decreased to 1: 1000 births. If there are antibodies in the blood of a pregnant woman against her child's red blood cells, their level is controlled additionally at week 28, 32 and 36. What is more, every 2 to 3 weeks, the doctor performs an ultrasound, which checks whether the child is developing properly. At low antibody values, no intervention is usually necessary, at very high pregnancy can be terminated earlier and the blood transfused into the child. In the “Rh-conflict”, intrauterine transfusion is possible. Red blood cells are transfused to foetuses in anaemia caused by the destruction of red blood cells by maternal antibodies (haemolytic disease of the foetus) also by parvovirus B19, also in the case of foetal bleeding into the mother's circulation, bleeding between monozygotic twins, and congenital defect of red blood cells, e.g. in thalassemia α. Transfusion in the newborn is usually made due to life-threatening hyperbilirubinemia, which cannot be reduced by phototherapy and is associated with foetal / neonatal haemolytic disease (CHHPN). Whole blood is used from a single donor or a preparation from a concentrate of red blood cells with fresh frozen plasma or a 5% albumin solution. Apart from the Rh antigen, other antigens responsible for blood groups (ABO, Kell, Kidd, Duffy) may be the cause of the serological incompatibility and haemolytic disease of the foetus and newborn. The conflict may also affect other blood cells, e.g. platelets (1: 1500 pregnancies) - then the antibodies produced by the mother lead to thrombocytopenia in the child or granulocytes (1: 6000 pregnancies), which results in granulocytopenia. Kell system The Kell system antigens are transmembrane glycoproteins with activity of zinc endopeptidase (with numerous SNP point mutations). The Kel gene locus is 7q33 and at least 36 unique antigens are associated with the Kell glycoprotein on erythrocytes - clinically relevant are: K, k, Kpa, Kpb, Kpc, Jsa, Jsb. The strongest immunogen is K antigen, which shows low frequency, but high immunogenicity. The antibodies anti-K (IgG) may cause haemolytic transfusion reaction, haemolytic disease of foetuses and newborn (destruction of foetal red blood cells and inhibition of erythropoiesis). Identification of group systems The basic method of identifying blood group antigens and antibodies against them is the agglutination reaction. Its occurrence depends on the number and location of antigens on the surface of the erythrocyte and the concentration of the corresponding antibodies in the serum. On the erythrocyte membrane, there are about a million molecules of the AB0 system antigens easily available for antibodies, because they are placed extracellularly, while the Rh system antigens are about 50,000 molecules placed intracellularly (intramembrane) more difficult to access, so it is more difficult to induce agglutination. For the detection of certain multi-sugar antigens, lectins are used - plant proteins that have the ability to bind sugars - antigenic determinants of some group systems. Examples of using lectins: 1. detection of AB0 system antigens, e.g.: - lectin from the plant Dolichos biflorus (seed of Asiatic plants of the legume family) – anti-A1, - lectin from Ulex europeaus - anti-H, 2. detection of MNS antigens, e.g. Vicia graminea - anti-N. Lectins from Dolichos biflorus are used to differentiate between the antigen A1 and its weaker forms, e.g. A2, which are less or not agglutinated in the presence of lectins. Lectins from D. biflorus bind α-N-acetyl-D-galactosamine (antigen A). Monoclonal antibodies The monoclonal antibodies are produced in vitro by the clone of cells - hybrids derived from the fusion of B lymphocytes (producing specific antibodies) and myeloma cells - tumour cells derived from a series of developmental B lymphocytes. They may be used to detect and determine the concentration of drugs, enzymes, hormones, in the diagnosis, localization and treatment of tumours, to obtain immunosuppression (e.g. after transplantation), to detect antigens of microorganisms, and as monoclonal reagents for the determination of blood group systems. Immunohematology - investigates the antigens found on the surface of blood cells and antibodies that are produced under their influence. Of more than 300 antigens found on the surface of red blood cells, only a few of them are of practical importance (e.g. AB0, Rh, Kell antigens). The common feature of blood cell antigens is that their character is inherited and transmitted in accordance with Mendel's laws, therefore they are used in genetic research. Blood transfusion Blood is a lifesaving liquid tissue. Whole blood is a mixture of cellular elements, colloids and crystalloids. As different blood components have different relative density, sediment rate and size, they can be separated when centrifugal force is applied. The components are prepared by centrifugation of one unit of whole blood. A single component required can also be collected by the apheresis procedure in blood donors. The apheresis blood donation is the process of blood collected via a special machine to separate it during the donation, so that only certain parts of the blood are collected and the remainder returned to the donor. Blood taken from a healthy donor can be separated during blood donation into its component parts. The needed component is collected and the "unused" components are returned to the donor. Fluid replacement is usually not needed in this type of collection. There are large categories of component collections: Plasmapheresis – blood plasma - for collecting FFP (fresh frozen plasma) of a particular ABO group; also immunoglobulin products, plasma derivatives, and collection of rare WBC and RBC antibodies may be received by the method. Erythrocytapheresis – red blood cells - the separation of erythrocytes from whole blood. It is most commonly accomplished using the method of centrifugal sedimentation. Plateletpheresis (thrombapheresis, thrombocytapheresis) – blood platelets - the collection of platelets by apheresis while returning the RBCs, WBCs, and component plasma. Leukapheresis – leukocytes (white blood cells) - is the removal of PMNs (polymorphonuclear leukocytes or polymorphonuclear neutrophils), basophils, eosinophils for transfusion into patients whose PMNs are ineffective or where traditional therapy has failed. There is limited data to suggest the benefit of granulocyte infusion. The complications of this procedure are the difficulty in collection and short shelf life (24 hours at 20 to 24 °C). Quality control demands that the product be irradiated to avoid graft-versus-host disease (inactivate lymphocytes). Irradiation does not affect PMN function. Since there is usually a small amount of RBCs collected, ABO compatibility should be employed when feasible. Stem cell harvesting – circulating bone marrow cells - harvested to use in bone marrow transplantation. The indications for administering blood preparations depend on many factors, such as the diagnosis of the disease, the stage, and type of treatment, the clinical condition and age of the patient. The decision to transfuse red blood cells should be made after all other treatment options have been exhausted. The European Committee on Blood Transfusion states: the clinical transfusion process is “the transfusion of the right unit of blood to the right patient at the right time, and in the right condition and according to appropriate guidelines”. It is a chain of inter-related events beginning with an appropriate decision that the patient needs transfusion of a blood component and ending with assessment of the clinical outcome of the transfusion (Guide to the preparation, use and quality assurance of blood components. Recommendation No. R (95) 15, 18th Edition European Directorate for the Quality of Medicines & HealthCare). https://www.avis.it/userfiles/file/News/EDQM%20Guide%2018th%20edition.pdf). Isolated blood components are currently used in blood transfusions, limiting the use of whole blood to a minimum. Blood collected from a healthy donor into a sterile plastic container with preserving fluid is split into components: red blood cells PRBC (packed red blood cells), platelets PLTC (platelet concentrate), plasma FFP (fresh frozen plasma), granulocytes, albumin solutions, immunoglobulins, clotting factors complex - coagulation factors. One unit is 450 ml of whole blood (± 10%), mixed with 63-70 ml of preservative liquid with anticoagulant for a final anticoagulant: blood ratio of approximately 1:7. It contains all the blood components that retain unchanged properties only for a certain time. Whole blood should not contain irregular antibodies of clinical significance. The obligatory serological tests include routine determination of ABO, Rh system groups and detection of irregular antibodies. There are two types of antibodies: natural (regular) or immune (irregular). The first of these - natural antibodies, occur in every human being, appear naturally in the first year after birth, and are present in the blood throughout their lives; they are antibodies to the main group antigens: anti-A (anti-A) and anti-B (anti-B). The second type of antibodies immune antibodies (irregular, are produced by the human body as a result of eliciting an immune response to foreign blood cells that have entered the circulation (e.g. after a blood transfusion). Recipients receive compatible blood in the ABO system. In addition, the Rh-negative recipient without D-antigen in blood cells can only receive Rh-negative blood. In all blood donors, a complete blood test for the ABO blood group and Rh antigen D as well as all other antigens is performed twice with blood samples taken at different times. Detection and identification of immune antibodies should be performed in all first-time and permanent donors who have been treated with blood during the year preceding the donation and in women with a prior history of pregnancy. Immediately before blood transfusion, the so-called cross-test - a serological compatibility test of the recipient and blood donor is made to confirm the compatibility of the blood of the donor and recipient in the main groups, i.e. in the ABO and Rh systems, and checked for the possible presence of antibodies to the blood donor's red blood cells in the blood of the recipient. The following tests are carried out during this test: 1) recipient and donor antigens are checked for AB0 group antigens; the presence of RhD antigen in the recipient is checked, and when it is RhD-negative, the D antigen in the donor is checked to avoid transfusion of RhD-positive blood cells to a RhD-negative person, 2) tests for the presence of immune antibodies in the recipient's serum, 3) a serological compatibility test between the recipient's serum and the donor's red blood cells. Serological compatibility of the donor and recipient is accepted when: 1) AB0 and RhD control confirms previous results of compatibility, 2) recipient serum does not react with donor blood cells (no agglutination), 3) testing for antibodies to the blood cell panel standards are negative or no additional antibodies detected but those previously identified. It should be noted that in patients requiring multiple transfusions for therapeutic reasons, new alloantibodies may occur (for antigens present on the surface of RBCs transfused). The serological compatibility test (cross-check) is valid for 48 hours from the time it is performed, because it is thought that during this time the immunization and production of antibodies in the recipient may occur. After this time, the cross test should be repeated. Incompatible transfusion (transfusion of blood components containing at least one antigen that is not present in the recipient) may involve blood cellular antigens as well as plasma protein antigens. In post-transfusion immunization, of greatest clinical significance are: 1. D antigen (Rh system), 2. K antigen (Kell system), 3. E, c, Cw antigens (Rh system) of D positive recipients. Complications of incompatible blood group transfusions occur on average every 3-5 thousand transfusions (early signs up to 24 hours after transfusion - 0.1% of patients and late - 1%). The causes are usually: transfusion of group blood prepared for another patient, replacement of blood samples for serological tests, or errors in serological documentation. As a result of such transfusion, haemolytic shock of varying severity may occur, and approximately twenty percent of patients suffer from hemorrhagic diathesis, which in about 50% of cases leads to death. The third most common complication is transfusion-related acute lung injury (TRALI), representing 7% of all transfusion complications (the incidence is estimated at 1: 2000 to 1: 7000 transfused blood units). Typical transfusion reactions in people without serious medical conditions are not fatal in 75% of the cases. For a long time, a universal donor was sought, a person whose biological material could be used for transfusion to any recipient. The erythrocytes of group Rh- O suspended in AB plasma (does not contain anti-A and anti-B antibodies) or suspended in plasma identical with the recipient group are considered universal blood. Scientists at the Scottish National Blood Transfusion Service have tried to obtain artificial blood from embryonic stem cells. Other scientists conducted experiments using bacteria, Streptococcus pneumoniae SP3-BS71, from which the enzyme glyosidic hydrolase was isolated – the enzyme responsible for cutting off the antigen end (N-acetylgalactosamine or galactose) from the RBC surface, forming a "universal blood cell"; however, the procedures used were not 100% effective. There are still a lot of tests to be done before clinical trials begin. Refferences: 1. Campbell, J.B. Reece: Biology. Pearson, Benjamin Cummings, Seventh Edition 2005 2. Jorde L.B.; Carey J.C.; Bamshad M.J.; White R.L.: Medical Genetics, Third Edition