ABO & Rh Discrepancies

1. ABO & Rh Discrepancies

2. Definition • When the results of the forward grouping (patient cells) do not correspond to the results of the reverse grouping (patient serum) or abnormal reactivity is present (i.e. Mixed Field) • i.e.theforward does NOT match the reverse

3. Patient A: Additional reaction with anti-B and patients cells. Patient B: Weak reaction with patients serum and A-cells. Patient C: Additional reaction with patients serum and A-cells. Patient D: Missing reactions with patients serum A-cells

4. ABO Discrepancies must be resolved • In RECIPIENTSthe discrepancies must be resolved before any blood component is transfused. • If not resolved before blood is needed, transfuse Group O (O NEGATIVE if there is a discrepancy in the Rh type also). • In DONORSthe discrepancies must be resolved before any blood is labeled with a blood type.

5. Categories of ABO Discrepancies • Cell Typing – Additional Reactions • Cell Typing – Missing Reactions • Serum Typing- Additional Reactions • Serum Typing- Missing Reactions

6. A- Cell Typing: Additional Reactions

7. 1- Polyagglutinable Red Cells • The removal of red cell N-Acetyl neuraminic acid by bacterial enzymes expose the T-Ag on the cell membrane. • Antibodies to T-antigens are naturally present in most human sera. • This Ab can also be found as a contaminant in some ABO typing reagents. • This cause unexpected agglutination of T Ags on red cells.

8. 2- Acquired Antigens • Microbial deacetylating enzymes such as E. coli cleave off the N-Acetyl of the Group A N-acetyl-galactosamine • The remaining galactosamine becomes similar to the Group B galactose • Anti-B react with this B-like Ag causing agglutination • A-like Ag can also be acquired

9. Acquired B Phenotype Group A individual N-acetyl galactosamine Galactosamine now resembles D-galactose (found in Group B) Bacterial enzyme removes acetyl group

10. 3- Non-specific Agglutination • Wharton’s Jelly – gelatinous substance derived from connective tissue that is found in cord blood and may cause false agglutination • Wharton’s Jelly Coats newborn cord cells and the child's type may appear AB.  • We do not do a reverse on newborn blood since they have not made any anti-A or anti-B yet.

11. If the baby types as an AB  recheck by washing cells several times and re-testing since you need to make sure you have removed the Wharton's Jelly and the baby is truly an AB.  • Better ALWAYS wash cord blood at least 4 to 5 before determining the type of the baby, or request new sample from heel

12. 4- Sensitized Red Blood Cells • Albumin in the ABO typing reagent can reduce the zeta potential • Effectively decreases the distance between red cells. • If the red cells are coated with antibody, false positive agglutination can occur

13. B- Cell Typing Missing Reactions

14. 1- Subgroups • Weak variants of both A & B • Carry poorly expressed Ags • May not produce expected reactions with anti-A & anti-B • They are categorized according to the strength and pattern of reaction with anti-A, anti-H & anti-A,B

15. 2- Altered Antigen Expression • Weak Ags may be found on RBCs of some people with diseases (Leukemia)

16. 3- Chimera • Chimera: Two cell populations • Natural Chimera: • In utero exchange of erythropoetic tissue between non-identical twins • Strength of reaction with ABO typing depends on the percentage of A or B cells in blood • Temporary Chimera: • following blood transfusion of ABO compatible, but not identical blood ( A received O cells)

17. 4- Excessive amounts of group specific substances • Patients with certain types of cancer • Large amounts of soluble A or B Ags • Inhibit anti-A or anti-B typing reagent • Can be resolved by washing RBCs prior to ABO typing

18. C- Serum Typing Additional Reactions

19. 1- Alloantibodies • Abs other than anti-A or anti-B • Can agglutinate A or B cells if express specific Ag • Abs commonly cause this discrepancy anti-M, -N, -S, -Lea, Leb, -P1, A1 • Can be identified by testing serum with a panel of O cells that have been phenotyped for these Ags

20. 2- Autoantibodies • IgM autoantibodies can cause false-positive results in cell & serum grouping • Problem can be solved by washing cells with warm saline prior to testing • In serum typing, autoabsorption can be performed • Or serum typing at 37oC

21. 3- Rouleaux formation • Rouleaux may also give false positive cell typing if strong enough • Looks like agglutination macroscopically • This phenomenon is due to alteration in serum protein concentration caused by: • elevated levels of gammaglobulins • elevated levels of fibrinogen • Also seen with plasma expanders (dextran) • Cell grouping- can be avoided by washing RBCs • Serum grouping- addition of saline

22. D- Serum Typing Missing Reactions

23. 1- Newborns • Infants develop ABO Abs by 3-6 months of age • Serum typing before this time: • Weak reaction • Negative reaction

24. 2- Patients with Hypogamma-globulinemia • Have low levels of immunoglobulins • Anti-A & anti-B may not be detected

25. 3- Chimera • Twins that have chimeric blood group can lack A & B Abs • Chimera with 98% O cells & 2% B cells • Group as O • Serum contain only anti-A

26. 4- Reagent Problems • Deterioration of Ags on A or B cells used for serum typing • Weak • Negative reaction

27. Rh Discrepancies • Rh –ve persons mistyped, & transfused with Rh +ve blood have 70% chance of becoming immunized • False +ve reactions can be identified by testing an Rh control with the patient’s red cells each time an Rh typing is performed • The test is not valid if the control causes agglutination

28. Causes of false positive reactions • Positive direct antiglobulin test • Polagglutinable red cells • Cold agglutinins or Rouleaux formation

29. 1- Positive direct antiglobulin test • The presence of Ab on patient’s red cells can cause a false +ve reaction with slide and tube anti-D • High protein medium reduces zeta potential allowing red cells to move closer • The cell bound Ab can cross link cells and cause agglutination

30. 2- Polagglutinable red cells • Rh –ve red cells that are polyagglutinable due to T or Tn activation • Agglutination occurs if anti-T or anti-Tn present in the anti-D reagent • Most anti-D reagents do not contain these antibodies

31. 3- Cold agglutinins or Rouleaux formation • Rh typing is performed using serum suspended red cells • If individual’s serum contains cold agglutinin or abnormal protein, false positive reactions can occur