Flow cytometric detection of platelet‐reactive antibodies and application in platelet crossmatching

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Abstract
Background: Alloimmunization against HLA or platelet antigens can cause refractoriness to platelet transfusions in multiply transfused patients. Crossmatching of platelet concentrates is effective in overcoming this problem. Study Design and Methods: A flow cytometric assay was used for simultaneous detection of lymphocyte‐reactive and platelet‐reactive antibodies in a single sample using fluorescein isothiocyanate‐labeled anti‐IgG. This assay was compared with the monoclonal antibody‐specific immobilization of platelet antigens (MAIPA) assay in selected sera containing HLA and platelet antibodies. In a further study, this assay was compared with lymphocytotoxicity test results from thrombocytopenic patients, for whom platelet concentrates were ordered. The results of both assays were then correlated with the 1‐hour corrected count increment, with a corrected count increment greater then 7500 considered as an adequate transfusion response. Results: The results of the MAIPA and flow cytometric assay in detecting platelet‐reactive antibodies correlated well (p<0.0001, r = 0.84). The sensitivity and specificity of the flow cytometric assay in detecting platelet‐reactive antibodies were 94.7 and 96.3 percent, when the MAIPA assay was taken as a reference. In unselected sera from patients, the sensitivity and specificity of the flow cytometric assays were, respectively, 72.7 and 91.7 percent in detecting lymphocyte‐ reactive antibodies and 70.6 and 77.7 percent in detecting platelet‐ reactive antibodies, when the lymphocytotoxicity test was used as a reference. With regard to an adequate transfusion response, the sensitivities and efficiencies were 20.0 and 82.1 percent, 33.3 and 84.3 percent, and 70.0 and 88.6 percent for the lymphocytotoxicity test and the lymphocyte‐reactive and platelet‐reactive flow cytometric assays, respectively. Conclusion: Flow cytometric crossmatching appears to be an effective method of detecting platelet‐reactive antibodies that may affect the success of platelet transfusions. This procedure is well‐suited for routine conditions and can be performed within 2 hours.