Blood Groups and Pre-transfusion Testing

• 1665: Dr. Lower successfully performs the first recorded blood transfusion between dogs, laying groundwork for future studies in transfusion medicine.

• 1667: A significant milestone occurs when a 15-year-old boy receives the first known animal-to-human blood transfusion, using blood from a sheep, highlighting early attempts to use animal blood for human treatment.

• 1818: Dr. Blundell, a pioneer in obstetric medicine, conducts a blood transfusion on a postnatal woman, utilizing her husband’s blood, marking a critical advancement in understanding human blood compatibility and transfusion practices.

• 1840: The first successful blood transfusion specifically aimed at treating hemophilia is performed, demonstrating the therapeutic potential of transfusions in managing specific medical conditions.

• 1901: Dr. Karl Landsteiner’s discovery of the ABO blood group system revolutionizes blood transfusions by categorizing blood types, reducing the risks associated with transfusions.

• 1914: The introduction of anti-coagulants significantly improves the safety and effectiveness of blood transfusions, allowing for better management of blood preservation during storage and transportation.

• 1930s: The establishment of the world’s first blood bank marks a pivotal development in the history of transfusion medicine, enabling systematic collection, testing, and storage of blood for future use.

• 1940s: The discovery of Rh blood groups and the establishment of the National Blood Transfusion Service enhance the understanding of blood group antigens and support more organized blood donation and transfusion efforts.

• 1960s: The introduction of anti-D immunoglobulin plays a crucial role in the prevention of Hemolytic Disease of the Fetus and Newborn (HDFN), providing critical care during pregnancy and protecting newborns from severe hemolytic conditions.

• 1980s: Blood donor screening practices are introduced, significantly reducing the transmission of infectious diseases through transfusions and improving the safety of the blood supply.

• 2005: The implementation of legislation and quality standards in blood transfusion practices ensures a systematic approach to blood safety and efficacy is upheld, promoting best practices across healthcare systems.

• 2017: Increased focus on avoiding unnecessary blood transfusions and exploring alternatives maximizes patient safety and outcomes, promoting the use of safer treatment options when possible.

What is Blood?

• Plasma: The non-cellular liquid component of blood, which transports essential proteins (e.g., Factor VIII), hormones, nutrients, and waste products, making it critical for maintaining bodily functions.

• Red Blood Cells (RBCs): Bioconvex, non-nucleated cells that contain hemoglobin, a protein responsible for the transport of oxygen from the lungs to the body's tissues and carbon dioxide from the tissues back to the lungs for exhalation.

• Platelets: Small, disc-shaped non-nucleated cells essential for blood clot formation; they play a crucial role in hemostasis by aggregating at injury sites and releasing chemicals that promote clotting.

• White Blood Cells: Various types of immune cells that act as key defenders against infections and diseases, with different functions including phagocytosis, antibody production, and immune regulation.

What is Blood Transfusion?

• Definition: The infusion of blood components or products into a patient’s circulatory system to restore or enhance blood volume and function.

• Components from donated blood: Blood can be separated into various components including whole blood, red blood cells, platelets, plasma, cryoprecipitate, and white cells, each tailored for specific medical needs.

Reasons for Blood Transfusion

• Patients often require transfusions due to various medical conditions that lead to a deficit in the production or maintenance of adequate blood components, including anemia, surgical blood loss, trauma, chronic diseases, and treatment side effects from chemotherapy.

Blood Components and Their Uses

• Red Blood Cells (RBCs): Commonly transfused for patients experiencing acute or chronic blood loss and severe anemia to restore oxygen transport capacity.

• Platelets: Administered for patients with low platelet counts (thrombocytopenia), frequently seen in conditions like leukemia or following chemotherapy.

• Fresh Frozen Plasma (FFP): Used to replace clotting factors in conditions such as liver disease, disseminated intravascular coagulation (DIC), or trauma-related bleeding.

• Cryoprecipitate: Provides high concentrations of fibrinogen and is utilized in cases of massive bleeding or for patients with coagulopathies.

• Granulocytes: Applied in severe sepsis cases when there is a need for increased neutrophil activity against infections.

• Albumin: Indicated for volume expansion in patients with burns, ascites, and significant liver disease, providing critical support in maintaining oncotic pressure.

Antigens and Antibodies

• Antigen: Any substance that can stimulate an immune response, leading to the production of antibodies in individuals lacking the specific antigen.

• Antibody: A type of immunoglobulin produced by the immune system in response to foreign antigens; essential for identifying and neutralizing pathogens.

• Blood Group Systems: Over 45 recognized blood group systems exist, featuring 362 red cell antigens as of November 2023. Understanding these systems is essential for safe blood transfusion practices.

The ABO Blood Group System

• Significance: The ABO system is the clinically significant blood group system based on the presence or absence of specific A and B antigens on red blood cells, which plays a critical role in transfusion medicine.

• Genetics: This system involves simple inheritance with three gene options (A, B, O), leading to four possible blood groups: A, B, AB, and O.

• Gene Dominance: A and B genes are dominant and co-dominant, while the O gene is recessive, indicating that individuals with type O blood lack A or B antigens entirely.

• Important Concepts:

  • O gene does not produce any phenotypic expression of antigens.

  • A and B alleles encode for enzymes (transferases) that modify a common precursor substance by attaching specific sugars to it, resulting in the distinct blood group phenotypes.

Inheritance Patterns

• Antigen Expression: The expression of A and B antigens arises from the addition of specific sugars to the H antigen by unique enzymes from parental gene contributions, leading to distinct blood types based on genotype combinations.

• Example Groups Include:

  • Group O: Only H antigen is present, no A or B antigens.

  • Group A: A antigen is present, determining the genotype as either AA or AO.

  • Group B: B antigen is present, determining the genotype as either BB or BO.

  • Group AB: Both A and B antigens are present, represented by the AB genotype, showcasing codominance.

Landsteiner's Law

• According to Landsteiner's discovery, when A or B antigens are absent, the corresponding antibodies are naturally produced in the serum or plasma, leading to potential reactions if incompatible blood is transfused.

• Antigens and corresponding antibodies are identified in the four blood groups: A, B, AB, and O, guiding clinical transfusion practices.

Rh Blood Group System

• Overview: Alongside the ABO system, the Rh blood group system includes over 50 antigens, with five major ones (D, C, c, E, e) that can influence blood transfusion compatibility and pregnancy.

• D Antigen: The D antigen is particularly crucial for transfusion; its presence indicates Rh-positive blood, while its absence results in Rh-negative blood phenotypes.

• D antigen is highly antigenic meaning it has strong reaction with its antibody

• D antigen is highly immunogenic meaning it is able to stimulate antibody production.

• RhD Gene: The RhD gene governs the expression of the D antigen, positioning it as a cornerstone of transfusion medicine.

Anti-D Prophylaxis

• D negative pregnant women are offered anti-D prophylaxis to prevent the production of maternal antibodies against the D antigen, protecting the fetus in cases of Rh incompatibility.

• This prophylactic treatment is typically administered at 28 weeks of gestation or following any sensitizing events during pregnancy.

Pre-Transfusion Testing

• Goal: The primary goal of pre-transfusion testing is to ensure safe transfusion practices, minimizing the risk of adverse reactions and optimizing blood management procedures.

• Testing Protocols: Testing protocols often include blood group typing, thorough antibody screenings, and crossmatching procedures to confirm compatibility between donor and recipient blood prior to transfusion.

Antibody Detection Methods

• A variety of techniques, including Direct Agglutination, Column Agglutination Technology (CAT), and Indirect Agglutination Test (IAT), are employed to identify blood group antibodies effectively.

• Crossmatching is a critical step, involving a compatibility assessment between the recipient’s plasma and donor red blood cells, ensuring safety in transfusion processes.

Conclusion

• A comprehensive understanding of blood groups, transfusion techniques, and underlying immunology is vital for ensuring patient safety and achieving optimal healthcare outcomes.

• Ongoing advancements in techniques, regulations, and clinical practices continue to support the evolution of safe and effective transfusion practices in modern medicine.

IgM and IgG antibodies are two important classes of immunoglobulins with distinct characteristics and functions in the immune response.

• IgM

  • Shape: Pentameric form, consisting of five monomer units linked together, giving it a large molecular size.

  • Placental Transfer: Unable to cross the placenta due to its size, which restricts its movement across biological membranes.

  • Red Cell Agglutination: Highly effective at causing agglutination of red blood cells due to its valency, which allows multiple binding sites to interact with antigens on RBCs.

• IgG

  • Shape: Monomeric form, consisting of a single unit, making it smaller than IgM.

  • Placental Transfer: Can cross the placenta, providing passive immunity to the fetus during pregnancy.

  • Red Cell Agglutination: Also capable of agglutinating red blood cells, but usually less effective than IgM due to its single binding site.

Understanding these differences is crucial in the context of blood transfusions and immune responses, particularly in conditions such as hemolytic disease of the fetus and newborn (HDFN), where maternal antibodies may impact fetal health.

H Antigen:

• Structure: The H antigen is a carbohydrate structure that serves as the foundation for the ABO blood group antigens. It consists of a fucose sugar attached to a galactose and N-acetylglucosamine molecule.

• Gene Product: The H antigen is synthesized by the action of the fucosyltransferase enzyme, encoded by the H gene. This antigen is present on the surface of red blood cells and is necessary for the formation of A and B antigens.

A Blood Antigen:

• Structure: The A antigen is formed by the addition of an N-acetylgalactosamine (GalNAc) sugar to the H antigen structure.

• Gene Product: The enzyme responsible for this addition is the A transferase, which is encoded by the A gene. This enzyme modifies the H antigen by attaching the N-acetylgalactosamine, resulting in the expression of type A blood.

B Blood Antigen:

• Structure: The B antigen is produced through the addition of a galactose sugar to the H antigen.

• Gene Product: The enzyme that facilitates this reaction is the B transferase, encoded by the B gene. The attachment of the galactose to the H antigen modifies it to express type B blood.

Important Concepts:

• The presence of either the A or B antigen (or both in the case of AB blood) is determined by the specific transferase enzyme activity from the corresponding gene, while the lack of these antigens corresponds to the O blood type where only the H antigen is present without any additional sugars.

Hemolytic Disease of the Newborn (HDN) occurs when there is an incompatibility between the blood types of the mother and the fetus, often related to the Rh blood group system. It typically arises when an Rh-negative mother carries an Rh-positive fetus. During pregnancy or delivery, if fetal red blood cells enter the maternal circulation, the mother's immune system may recognize the Rh-positive cells as foreign and produce anti-D antibodies in response.

These antibodies usually IgG class, can cross the placenta in subsequent pregnancies, attacking the fetal red blood cells, leading to hemolysis (destruction of red blood cells), anemia, jaundice, and potentially serious complications for the newborn, such as heart failure and developmental issues. This condition underscores the importance of anti-D prophylaxis for Rh-negative pregnant women to prevent the sensitization to the D antigen.

Additional Blood Group Systems

Kell Blood Group System

• The Kell blood group system includes over 20 antigens, with K (Kell) being the most clinically significant.

• Anti-K can cause hemolytic transfusion reactions and hemolytic disease of the newborn.

• K antigen is highly immunogenic, meaning it can provoke an immune response.

Duffy Blood Group System

• The Duffy blood group system consists of two main antigens: Fya and Fyb.

• These antigens serve as receptors for malaria parasites (specifically Plasmodium vivax), influencing susceptibility to malaria.

• Individuals with Fy(a-b-) phenotype are resistant to P. vivax.

Kidd Blood Group System

• The Kidd blood group system comprises antigens Jka and Jkb.

• Antibodies against Kidd antigens can cause delayed hemolytic transfusion reactions.

• These antibodies are often difficult to detect due to their low levels in serum.

MNS Blood Group System

• The MNS blood group system is defined by the presence of M, N, and S antigens on red blood cells.

• This system includes multiple antigens, which can have implications in transfusion medicine and hemolytic disease.

• Variants in these antigens can affect compatibility and immune response during transfusions.

Lewis Blood Group System

• The Lewis blood group system consists primarily of Lea and Leb antigens, which are not intrinsic to red blood cells but are instead found in saliva and other tissues.

• Lewis antigens are formed by the action of specific enzymes and their expression can change during pregnancy.

• Anti-Lea and anti-Leb antibodies are generally considered less clinically significant, although they can still cause complications.

Lutheran Blood Group System

• The Lutheran blood group system has two main antigens: Lu(a) and Lu(b).

• The expression of these antigens can vary with the patient's age and health status.

• Antibodies against Lutheran antigens can be clinically significant but are relatively rare.

Understanding these blood group systems is crucial for ensuring safe blood transfusion practices and managing potential hemolytic reactions, especially in settings involving pregnancy or complex transfusion scenarios.

Direct Agglutination:

• Direct agglutination involves the clumping of cells or particles that possess specific antigens when mixed with antibodies in a solution. This technique typically uses a liquid phase where the antibody interacts with the antigen directly on the surface of the cells (e.g., red blood cells).

• Solid phase is not employed in this method as the reaction occurs free in solution.

Column Agglutination Technology (CAT):

• Column agglutination technology (CAT) is an advanced method that utilizes a solid-phase support matrix (often in a column format).

• In this method, antibodies are immobilized on the solid phase, allowing for the binding of any specific antigens present on cells in a liquid phase that flows through the column.

• This enables a sensitive detection of agglutination as the antigen-antibody complexes can be easily visualized and measured as they remain adhered to the solid phase.

Indirect Agglutination:

• Indirect agglutination involves the use of coated particles, such as red blood cells or latex beads, that are bound to antibodies in a solid phase. The liquid phase containing antigens will cause these coated particles to agglutinate, forming visible clumps.

• This method combines elements of both phases as the antibody is attached to a solid phase particle while the antigen is in a separate liquid phase.
  Understanding these methods is crucial for effective diagnostics and blood typing practices.

Indirect Antiglobulin Test (IAT) using Anti-Human Globulin (AHG) cards is a laboratory method utilized to detect antibodies against specific blood group antigens in a patient’s serum. The process generally involves the following steps:

1. Sample Preparation:

   • A patient’s serum is collected and potentially mixed with red blood cells (RBCs) that have known antigens.

2. Incubation:

   • The serum and RBCs are incubated together to allow any antibodies present in the serum to bind to the corresponding antigens on the RBCs.

3. Washing:

   • After incubation, the cells are washed to remove unbound antibodies, reducing background and ensuring a clear result.

4. AHG Addition:

   • An AHG reagent is added, which binds to any IgG antibodies that are attached to the RBCs. This step is crucial as it facilitates the detection of bound antibodies.

5. Agglutination Read:

   • The mixture is assessed for visible agglutination. If agglutination is observed, it indicates that the patient’s serum contained antibodies against the antigens present on the tested RBCs.

6. Card Format:

   • AHG cards streamline this process, containing wells or chambers pre-filled with specific reagents, which simplifies and standardizes the testing procedure. The cards enhance ease of use and interpretation of results while minimizing contamination risks.

IAT is important for blood typing, crossmatching prior to transfusions, and prenatal testing to identify potential hemolytic disease of the newborn (HDN).

Antibody Screening: A laboratory procedure used to identify the presence of antibodies against specific blood group antigens in a patient’s serum. This is crucial in transfusion medicine to ensure compatibility between donor and recipient blood. The process typically involves mixing the patient’s serum with red blood cells of known antigens to observe any agglutination reaction, indicating the presence of corresponding antibodies.

Crossmatching: A critical step in the transfusion process, crossmatching involves testing a sample of the recipient's plasma against donor red blood cells to assess compatibility before transfusion. It ensures that the donor blood will not cause an adverse reaction in the recipient's immune system. The procedure can include both immediate spin crossmatch (for ABO compatibility) and further compatibility testing using methods such as indirect antiglobulin testing (IAT) for better antibody detection and potential incompatibilities.