[Molecular blood genotyping in patients with Thalassemia major in Tehran adult thalassemic clinic]

Blood typing by serologic methods after transfusion has limitations due to presence of donor red cells in recipients. Accurate determination of red blood cells [RBCs] antigens is very important in multitransfused patients including beta-thalassemics and sickle cell anemics. So, the aim of this study was to evaluate DNA- based methods as supplement to the hemagglutination technique to determine the red blood cell [RBC] antigen profile of multitransfused patients with beta- thalassemia. DNA was extracted from peripheral blood of 20 apparently normal people and 44 patients including 35 with beta- thalassemia [out of whom 19 had clinical evidence of delayed hemolytic transfusion reaction], 8 with thalassemia intermedia [out of whom 2 had hemolytic reaction], and one with sickle cell thalassemia. RHD/ RHC/ RHc/ RHE/ RHe/ JKA/ JKB/ FYA/ FYB/ KELL1/ KELL2 alleles were determined by PCR and were then compared with the hemagglutination method. Phenotype and genotype results were the same in all controls. The phenotypes and genotypes of 53 blood antigens of 26 patients were incompatible. Most of the discrepancies [19 cases] occurred in the Rh system, and fifteen in the Duffy and Kidd systems. The results show that screening platelet concentrates for bacterial contamination is necessary for blood transfusion centers and hospital blood banks. Blood typing by serologic method was not accurate in this study but genotyping could determine true blood groups in multitransfused patients and help in selection of RBCs without alloimmunized antigens in future transfusion attempts. Specificity, sensitivity, positive and negative predictive values of hemagglutination method for RhD antigen had good values in comparison to the molecular method. This might be due to pre- transfusion determination of RhD for thalassemic patients so as to receive Rh- matched blood units. It seems pre-transfusion blood typing of Rh and Kell antigens, which are the cause of hemolytic reactions, in comparison to the molecular method could be cost effective. In addition, typing of Rh and Kell antigens in some regular blood donors could be helpdul for selecting antigen-negative RBCs for transfusion dependent patients

[Flow cytometric survey of RBC chimerism after bone marrow transplantation]

The aim of the present study was to evaluate red blood cell chimerism after bone marrow transplantation by flow cytometry. In order to perform this assay, FITC labeled antibodies against blood groups ABH, Rh, Kell, Duffy, Kidd, MNS were used.14 hematologic patients under BMT were selected for this study. The required sample was 5 ml peripheral blood that is collected in tubes containing EDTA. At first, donor and recipients red cells phenotypes were identified with the use of both agglutination and flow cytometry methods; then, on post-transplantation days of 15, 30 and 60, only blood samples of the recipients were analyzed by flow cytometry for the antigens differing from donors to recipients. Antibody screening test and titration of ABH Isohemagglutinins were performed on recipients' plasma samples and then repeated on post-transplantation day of 60. After BMT, red cell chimerism was detected in all 14 patients [in 9 patients on post-transplantation day of 15 and in 5 patients on day of 30]. Antibodies against minor blood groups and Rh blood group were not detected at all. The occurrence of chimerism was not inhibited by ABO incompatibility of donors and recipients but in patients who were ABH incompatible with their donors, ABH isohemagglutinins titer following transplantation decreased. Although the presence of isohemagglutinins did not prevent chimerism but it seems these antibodies by attaching to their related antigens on chimeric red cells membrane prevented corresponding antigen detection. Now by using flow cytometry, red cell phenotyping is applicable and reticulocyte analysis is much easier to perform so that chimerism can be detected in patients who have recently experienced blood transfusion. Moreover, through further evaluation of red cell chimerism and detection of recipient autologous red cells, disease relapse can be predicted