Blood grouping and cross matching practi
- 1. Blood grouping and Cross matching, Compatibility testing and Transfusion reactions Dr sonal meshram
- 2. Blood groups The term is applied to genetically determined antigens that can be detected on the surface of red cells by the specific antibodies Karl Landsteiner -- Nobel prize in 1930
- 3. Landsteiner law 1. If an agglutinogen is present on red blood cell membrane, the corresponding agglutinin must be absent in the plasma. 2. If an agglutinogen is absent on red blood cell membrane, then corresponding agglutinin must be present in the plasma
- 4. Blood group antigens designated by capital letters, small letters, name of discoverer ISBT recognizes 30 blood group systems
- 5. Genetic loci of blood group antigens locus for ABO group - chromosome 9 Rh group - chromosome 1 H group - chromosome 19
- 6. Types of blood group Antibodies Naturally occurring Abs Immune -derived Abs predominant Ig M type complete abs predominant IgG incomplete abs. Capable of agglutinating red cells in saline suspension can not agglutinate
- 7. AB antibodies are naturally occurring Ig M antibodies and react with strongly at temperatures lower than 37C. Anti- A and Anti - B present in newborn at birth are passively acquired from mother in- utero child begins to develop own Abs at 3-6 months age as gets exposed to antigenically similar antigens
- 8. Rh system Rhesus monkey red cells injected into rabbits and guinea pigs antibody which was raised reacts with rhesus monkey red cells as well as with 85% human red cells • Antigen involved was called as Rh factor
- 9. Antigens of Rh system • C, D, E, c, e. • D antigen is the most immunogenic. • Depending on presence /absence of D antigen on red cells Rh positive ( 95%) Rh negative(05%)
- 11. Blood grouping - ABO grouping • Two methods - 1) Cell grouping (forward grouping): Red cells are tested for the presence of A and B antigens employing known specific anti-A and anti-B (and sometimes anti-A, B) sera. 2) Serum grouping (reverse grouping): Serum is tested for the presence of anti-A and anti-B antibodies by employing known group red cells
- 12. Cell grouping (forward grouping) 5 methods • Slide test • Tube method • Microplate method • Column agglutination or gel technology • Solid phase adherence technology
- 13. Slide test • Anti-A is colour coded blue • anti-B is colour coded yellow.
- 14. Principle • Red cells from the specimen are reacted with reagent antisera (anti-A and anti-B). Agglutination of red cells indicates presence of corresponding antigen (agglutinogen) on red cells. Specimen • Capillary blood from finger prick, or venous blood collected in EDTA anticoagulant. Reagents • ABO antisera - monoclonal
- 16. Advantages of Slide test: rapid and simple. Disadvantages of slide test : • Weakly reactive antigens (like A2) may be missed by slide method. • Slide test is also not suitable for serum grouping. • Drying • If the result is read before two minutes, weak antigens may be missed. Results of slide test should always be confirmed by cell and serum grouping by tube method.
- 17. Tube method For cell grouping/forward blood grouping: • known antiserum and patient’s RBCs are mixed in a test tube, • incubated at room temperature for 5 minutes, • and then centrifuged. • Following centrifugation, a red cell button (sediment) will be seen at the bottom of the tube. • Cell button is resuspended by gently tapping the base of the tube
- 19. Positive and negative controls Advantages of tube method: more reliable than slide method as centrifugation brings antigen and antibodies closer together & allows detection of weaker antigen antibody reactions
- 20. M i c r o p l a t e m e t h o d • A microwell plate consists of 96 small wells (U- or V- bottom shaped). • U-type is preferable since results are easier to read.
- 21. DONOR SELECTION
- 22. Medical history - Age - Donation interval - Volume - Pregnancy and lactation - Infectious diseases - Illness - Drugs - Immunisation
- 23. Physical examination • Weight • BP • Pulse • Temperature
- 24. • Hb estimation by copper sulphate method Specific gravity of 1.053 is equivalent to Hb concentration of 12.5 gm/dl
- 26. Blood components and derivatives
- 27. Platelet incubator Refrigerated centrifuge Refrigerators
- 29. Cross matching / Compatibility testing : • Consists of : -testing patient’s serum against donor RBCs to detect any antibodies in patient’s serum, reacting with donor RBCs (major cross match) -testing patient’s RBCs against donor serum to detect any donor antibodies reacting with patient’s RBCs (minor cross match). • Donor blood is considered to be compatible if there is no agglutination or haemolysis in major or minor cross match.
- 30. • Detect any irregular antibodies present in recipient’s serum reactive against donor’s RBCs -Those antibodies other than anti-A and anti-B in recipient’s serum are called as irregular or unexpected antibodies.
- 31. TESTS/ METHODS OF CROSS MATCHING: • A single tube cross match consisting of three stages is recommended. • Recipient’s serum is tested against donor’s RBCs under three different conditions; • Agglutination or haemolysis in any one of the three stages indicates incompatibility. • The three stages of compatibility test are as follows: 1. Compatibility test at room temperature 2. Compatibility test at 37° C 3. Indirect antiglobulin test (Indirect Coomb’s Test)
- 32. spi n cros s 1. Compatibility test at room temperature (Immediate match): • The purpose of this test is to detect ABO incompatibility. • Saline-suspended donor’s RBCs and recipient’s serum are mixed in a test tube, incubated briefly at room temperature, and centrifuged. • Agglutination or haemolysis indicates incompatibility. • If agglutination or haemolysis is absent, next step is performed.
- 33. 2. Compatibility test at 37°C: • The test tube from test performed at room temp. ( ie. Test tube from stage 1) is: - incubated at 37° C for 20 minutes, - centrifuged again, and - examined for agglutination. •Agglutination or haemolysis indicates incompatibility • Ifagglutination is absent, perform the last stage (ie. stage 3).
- 34. 3. Indirect antiglobulin test (Indirect Coomb’s Test): • The mixture from stage 2 is: - incubated at 37° C for 30-60 minutes, - washed in saline, - antiglobulin reagent (Coomb’s sera) is added. - re-centrifugation, - examine for agglutination or haemolysis. • This test detects most of the clinically significant IgG antibodies. • If agglutination or haemolysis is not observed in any of the above stages, donor unit is considered to be compatible with recipient’s serum.
- 36. Immediate Immunological • FNHTR • Haemolytic transfusion reaction • Allergic reactions • Anaphylactic reaction • TRALI Non- Immunological • Circulatory overload • Bacterial contamination
- 37. DELAYED Immunological • Hemolytic transfusion reaction • GVHD • Post transfusion purpura Non-Immunological • Transmission of infectious organism • Iron overload
- 38. THANK YOU