The detection of platelet alloantibodies by flow cytometry

A flow cytometric procedure was investigated for its ability to detect antibodies directed against blood group A, HLA, and PlA1 (HPA-1a) antigens. When type O sera were tested against platelets from blood group A donors, only 9 of 14 positive reactions were observed. Furthermore, the expression of blood group A varied more than 100-fold on platelets derived from individual donors. When anti-HLA-A2 and -B7 were evaluated, 11 of 11 individuals with HLA-A2 and -B7 antigens reacted. In contrast, when platelets from donors whose HLA antigens included HLA-B8 or -B12 were tested with anti-HLA-B8 or -HLA-B12, respectively, positive reactions were observed in only 3 of 7 instances, despite the fact that the lymphocytes reacted strongly. Platelets from 10 HLA-A2-positive donors, which had been stored for up to 20 months at -70 degrees C, were studied. In all cases, frozen-stored platelets reacted well with an anti-HLA-A2. Limited testing with an anti-PlA1 (anti-HPA-1a) showed equal reactivity with fresh and frozen platelets. Finally, the method was compared to a visual immunofluorescence assay using sera from patients who were refractory to platelet transfusions. The results agreed in 30 of 37 comparisons, and most discrepancies were resolved in favor of flow cytometry. It is concluded that flow cytometry is useful for detecting platelet alloantibodies and possibly for prospective platelet crossmatching, as HLA- and platelet-specific antibodies can be identified by using platelets stored frozen for several months.