1 / 46

Immunohematology Transfusion Medicine Blood Bank

Immunohematology Transfusion Medicine Blood Bank. History 101 of Blood Bank. 1492: first attempt at using blood for therapeutic use. 1665: First animal to animal-Richard Lower 1667: Jean Baptiste Denys, first successful IV transfusion of blood from animal to human

bernad
Télécharger la présentation

Immunohematology Transfusion Medicine Blood Bank

An Image/Link below is provided (as is) to download presentation Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author. Content is provided to you AS IS for your information and personal use only. Download presentation by click this link. While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server. During download, if you can't get a presentation, the file might be deleted by the publisher.

E N D

Presentation Transcript


  1. ImmunohematologyTransfusion MedicineBlood Bank History 101 of Blood Bank

  2. 1492: first attempt at using blood for therapeutic use. • 1665: First animal to animal-Richard Lower • 1667: Jean Baptiste Denys, first successful IV transfusion of blood from animal to human • 1818: James Blundell First to transfuse human to human. • 1901: Karl Landsteiner: Discovered ABO blood grouping

  3. 1927: Landsteiner and Levine discovered M,N and P system • 1939,40: Levine, Stetson, Landsteiner and Weiner discovers Rh system and it’s role in erythroblastosis foctalis (HDN) • 1946-Kell system discovered by Coombs, Mourant and Race • 1950-51: Duffy, Kidd, Lutheran system discovered. • Landsteiner and Alexanders lead to the discovery of >800 Blood group systems.

  4. Antigen and Antibody in blood banking • Antibody production is result of blood group antigen (foreign-transfusion or pregnancy), or can be naturally occurring. • Blood group antigens are integral part of the RBC membrane • B Cells produce antibody molecules that are specific for a target antigen (part of the surface of RBC) • Antigenic determinants: on RBC-elicit the production of different antibodies

  5. Complement System: set or group of serum proteins that generates membrane attack that causes cell destruction. (Hemolysis) • Classical vs Alternative pathway • Antibody-Antigen complex activates Complement. • Complement activation • Anaphylatoxins • Vasoactive amines • Chemotactic • Opsonins • Receptors

  6. Antigens-have different levels of immogenicity-cause immune response. • Factors effecting immunogenecity • Chemical composition and complexity • Degree of foreignness • Size • Dosage • Route of administration

  7. Red Blood Cell Antigens • 23 blood group systems with more than 200 RBC antigens • RBC antigens are determined by genetic inheritance pattern • Antigen immunogenicity based on stimulation production upon exposure • Types of antigen RBC, HLA, Platelet

  8. Antibodies-protein (immunoglobulin-Ig) • 5 Classification: • IgG- Clinically significant in BB • IgM- Can be Clinically significant in BB • IgA • IgE • IgD

  9. IgM antibodies: • Produced initially in response to foreign antigen • Large pentamer structure • Contains 10 potential antigenic sites • In BB – reaction in saline procedures • 5-10 % of Ig class • Short half-life 5-6 days • Activates Complement with great efficiency.

  10. IgG antibodies • Monomer • Accounts for 80% of Ig • Found in extravascular fluid • 2 antigen binding sites • In BB- reaction less apparent in saline procedures, need additive to show reaction. • Half-life 23 days • Cross placenta • Activates complement • Subclasses

  11. Immune response (Primary vs. Secondary) • Immune response influenced by: • Age • Route of exposure • Genetic make-up • Health

  12. Antigen-Antibody interaction: • Formation of antigen –antibody complex causes an immune reaction. • Amount is determined by: • Goodness to fit • Size • Shape • Charge

  13. Antigen-Antibody reaction in Vivo vs. Vitro • Primary concern in Blood Bank-Vivo • Want to reduce the potential of exposure to foreign antigens which would result in antibody production. • Identify antigens present on the RBC • Identify any antibodies produced – found in the serum/plasma

  14. Identify antigen-antibody complex by: • Hemolysis (Complement reaction associated with complex) • Agglutination (antigen/antibody complex), like clotting of the blood Don’t want agglutination- 2 stages of agglutination: • Sensitization- antibody binds to an antigen. Is influenced by amount of antigen and antibody present.

  15. Sensitization enhanced by: • Serum to cell ratio • Temperature • Incubation • pH • Ionic strength

  16. Lattice Formation stage (cell interaction, see visible agglutination, linkage between antigen and antibody) • Factors that influence Lattice formation • Zeta potential • Serum to cell ratio (zone of equivalence) • Prozone effect • Centrifugation Grading agglutination (lattice formation) 0(neg), 1+, 2+, 3+, 4+

  17. Indicators of antibody-antigen complex • Hemolysis • Agglutination Hemolysis is an indication of antigen-antibody complex that results in cell destruction. Identified usually in the Antiglobulin Test.

  18. Antiglobulin Test (Anti-humanglobulin or AHG) • Discovered by Coombs, Mourant and Race in 1945- found that RBC can become sensitized without visible agglutination. • Reagent base test • Identifies IgG antibodies and Complement proteins • Polyspecific and monospecific AHG reagent.

  19. AHG reagent will react with bound and/or free IgG and Complement proteins. • Must get rid of the free to I.D. the bound forms that can cause reaction. • WASH cells (RBC for testing-cell suspension) • If cells wash properly and add AHG reagent and agglutination forms this is a positive reaction (NOT GOOD) • 2 Types of AHG test: • Direct antiglobulin test (DAT) • Indirect Antiglobulin test (IAT or antibody screen)

  20. Direct antiglobulin Test: • Detects antibodies bound to RBC in vivo • Results in clinical event or illness • (+) DAT indicates an immune response; patients cells have attached IgG and/or Complement) • EDTA is sample choice for DAT

  21. Indirect Antiglobulin Test (IAT) • Detects in vitro sensitization of RBC • 2 Step process: • Incubation at 37°C serum with donor cells False positive and False negative reaction for various reasons. Positive IAT indicates a specific reaction between antigen and antibody in serum of patient.

  22. Potentiator Reagent: enhances antibody and antigen complex • A.K.A. as enhancement media • Enhances antibody uptake • Promotes agglutination • 4 Types potentiators • Low ionic strength solution (LISS) • Bovine serum albumin • Polyethylene glycol (PEG) • Proteolytic enzyme (3 Types) • Papain • Ficin • Bromelin

  23. Chapter 2: Blood Bank Reagents • Basic blood banking reagents depends on the source of antigens and source of antibody in testing. (What are we looking for?) • Detect an antigen present or absent on RBC (donor/patient cells) • Detect antibody present or absent in serum (donor/patient serum) Have to have a known source of antigen to detect antibody or known antibody to detect antigen.

  24. Source of Antigens: • Commercially prepared RBC suspension with known antigens to detect unknown antibodies. • In BB- usually patient RBC antigen type is unknown and we test the cells to identify the type of antigen present on the RBC by testing with a known antibody • I.D. blood type (forward typing)

  25. Source of Antibody • Anti-sera commercially prepared or can be patient serum or plasma in BB. • Commercially prepared anti-sera contain known RBC antibodies to identify unknown RBC antigens. • Reverse Typing

  26. Routine Testing procedure: • ABO/Rh typing • Antibody screen (IAT) • Antibody Identification • Cross match Only do antibody identification if IAT is positive. If IAT is negative-indicates there are no unknown antibodies present in patients sample.

  27. Important to understand reagent so that you understand what you are looking for or identifying Antigen Antibody ABO/Rh Pt. RBC Commercial Anti-A, Anti-B, Anti-D Antibody Screen Screen cells Pt. Serum Antibody I.D. Panel Cells Pt. Serum Cross-match Donor cells Pt. Serum

  28. Reagents – 4 basic groups • Reagent RBC- Known RBC antigens • Antisera- Known RBC antibodies • Antiglobulin (AHG) • Potentiators Reagents are regulated by the FDA: minimum standards for reagents used for testing. • Specificity • Potency

  29. Reagent Quality Control • Technical procedures to determine analytical testing phase work properly. • Specific Q.C. requirements • Checks: • Reagents • Equipment

  30. Reagents are: • Monoclonal (single clone of cells- specificity) • Advantage vs disadvantage 2. Polyclonal (human source: mixture of cells-contains multiple antibodies) Assist in identifying antigen present on patients RBC that may cause and reaction in vivo.

  31. Anti-Sera for ABO typing (forward typing): • Anti-A and Anti-B determines the presence of A,B or no antigens on RBC. • Test performed using Donor/Patient RBC with known Anti-sera • (+) agglutination = antigen present • (=) agglutination = antigen absent • Identifies the 4 major blood groups A: posses A antigens B: posses B antigens AB: posses both A and B antigens O: lack antigens (has no antigens)

  32. Anti-sera for Rh typing: • There are multiple antigens in the Rh system (c,C,D,E,e), most prevalent and most important to identify is D. • Identify the presence (+) or absence (=) of the D antigen on patient RBC. • Use anti-D reagent combined with testing RBC. • Agglutination: D present, D (+) • No agglutination: D absent, D(=)

  33. Antiglobulin Reagent: (AHG) • Detects IgG antibodies and Complement protein that have attached to RBC. • 2 Types • Polyspecific • Monospecific

  34. Check Cells (Coomb’s Control Cells) • Required control system by AABB standards • Control system: • RBC commercially prepared with IgG antibodies • Identified true negative reactions-false negative

  35. Reagent RBC • Reagent A1- RBC with known antigen (A) mixed with serum to identify or confirm ABO typing • Reagent B- RBC with known antigen (B) mixed with serum to identify or confirm ABO typing • Testing phase known as Reverse typing or Back typing.

  36. Patient posses the antibody directed against the antigen of their ABO group. • Example: Group A individual: A antigens on RBC; Lack B antigen; produce B antibodies (in serum or plasma). Serum agglutinates with B cell reagent. • Group B individual: B antigens on RBC; Lack A antigen; produce A antibodies (in serum or plasma). Serum agglutinates with A cell reagent

  37. Screen Cells-Group of known reagents that contain known antibodies. • Looks for antibodies with specificity to RBC antigens in patient and donor sample (naturally occurring or from exposure) • Commercially available: Group O donor source • AABB standards states antibodies test performed on recipient specimen required • Blood group antigens expressed on or in screen cells: D, C, E, c, e, M, N, S, s, P1, Lewis, Lutheran, Kell, Duffy and Jka, Jkb

  38. Panel Cells (10 + reagent based testing system) • Group of test to determine the specificity of RBC antibody that was Identified in antibody screen. • Antigenic profile is important. • Lectins which are useful in identifying certain antibodies through panel cells. • Lectins: pg 50

  39. Testing procedure uses tube method and slide method. • New method include: • Gel technology • Microplate • Solid phase – serological method-automative

  40. Chapter 3: Genetics • Blood type is determined by genetic inherited patterns. • Phenotype: observable trait • Genotype: actual genetic make-up • Predict genotype, if you know phenotype and can predict phenotype, if you know genotype. • Blood type is determined by the antigen present on the RBC. • Punnet Square

  41. Genes: unit of inheritance on a chromosome. They are located on specific areas of the cells called genetic loci. • Alleles: Form or different forms of a gene of a given loci • Ex: A, B and O alleles on the ABO gene locus. • Polymorphic: having two or more alleles at a given locus. (Rh system)

  42. Inheritance Pattern: • Co-dominant (equal expression of a gene on an individual) • Recessive or dominant ( only one alleles is expressed on the cell) • Amorphic expression: gene present, but does not express detectable product. Mandelian Principle: Independent segregation of traits 4. Mutations

  43. Homozygous vs Heterozygous inheritance • Homozygous: 2 alleles for a given trait are the same-genotype are identical genes • Heterozygous: 2 alleles are inherited are different-genotype are different. • Agglutination reactions will be effected by inheritance pattern. • Homozygous pattern- stronger reactions • Heterozygous pattern- weaker reactions

  44. Inheritance pattern with Cis and Trans position. (related to the Rh system and how it expresses itself) • Cis : gene expression is from the same chromosome • Trans: gene expression is from different or opposite Chromosome. • Can help determine agglutination reaction.

  45. Silent Genes- “Amorph”, present and cause problems, but do not produce detectable antigen. • Result in unusual phenotype • Example is a “Null” type individual- blood type is not apparent or predictable- see with the Rh system.

  46. Paternity Testing • Direct exclusion: child has a trait present that neither parent posses. • Indirect exclusion: child lacks a gene that should be inherited from the parent in question. • Inclusion: when the child has the predictable traits that are expected form the parent in question.

More Related