Development of quantitative monoclonal antibody-specific immobilization of platelet antigens assay for antibodies against human platelet antigen-1a, 3a,and 5b

Human platelet antigens (HPAs) are platelet-specific alloantigens that cause alloimmunization. Alloantibodies induced via pregnancy and transfusion may result in various clinical syndromes. The monoclonal antibody-specific immobilization of platelet antigens (MAIPA) assay is the gold standard for the detection and identification of HPA antibodies. The present study aimed to develop standardized quantitative MAIPA assays for antibodies against HPA-1a, 3a, and 5b. MAIPA protocol was applied on serial dilutions of standard or reference reagents containing antibodies against HPA-1a, 3a, and 5b. Optimal experimental parameters were determined, and standard curves were constructed with a linear regression. The sensitivity, specificity, and reproducibility of the assays were also evaluated. The optimum reagents and conditions for the MAIPA assays were decided through repeated testing and comparison. The intra-assay coefficients of variation (CVs) are 1.3~3.6%, 2.7~5.2%, and 4.7~5.3%; the inter-assay CVs are 4.0~5.6%, 4.4~5.9%, and 3.2~6.3%, respectively, for HPA-1a, HPA-3a, and HPA-5b antibodies. In specificity tests, some monoclonal antibodies and sera with other antibodies were tested and no obvious positive result was observed. The standardized quantitative MAIPA assays for antibodies against HPA-1a, 3a, and 5b have good sensitivity, reproducibility, and specificity. They have important application value in clinical diagnosis of alloimmunization disease caused by HPA antibodies and safe transfusion of platelets.