Platelet antibody detection assays: a single-laboratory comparison of MAIPA, PIFT, and microsphere-based multiplex assays Pak-Lx

Background and Objectives: The identification of platelet antibodies is essential for diagnosing and managing conditions such as fetal and neonatal alloimmune thrombocytopenic purpura, post-transfusion purpura, and immune platelet refractoriness. Monoclonal antibody immobilization of platelet antigens (MAIPA) is the standard method for detecting anti-human platelet antigen (HPA) antibodies, while the detection of anti-HLA antibodies once relied on the complement-dependent cytotoxicity method, however advanced technologies such as enzyme-linked immunosorbent assay and Luminex have significantly improved sensitivity and accuracy in identifying these antibodies. Flow cytometry-based techniques (platelet immunofluorescence test - PIFT) and Luminex platform-driven microsphere-based multiplex assays (Pak-Lx) are widely employed in platelet immunology laboratories owing to their remarkable flexibility and versatility. The present study compared the sensitivity, specificity, and concordance of these different serological techniques used in platelet antibody identification. Material and Methods: One hundred serum samples from patients suspected of immune-mediated platelet disorders were examined. Initially, the samples underwent testing using the MAIPA method. Subsequently, the results were compared with three alternative methods: PIFT and microsphere-based multiplex assays for both HLA and HPA antibodies. Results: Pak-Lx demonstrated a 94 % agreement with MAIPA, while PIFT had 88 % agreement for HPA antibodies. For HLA antibody detection, Pak-Lx versus DLX had 75 % concordance, MAIPA versus DLX showed 77 %, and PIFT versus DLX displayed an 81 % concordance rate. Remarkably, there were no significant differences in concordance levels between Pak-Lx and PIFT compared to MAIPA and DLX for anti-HPA and HLA antibodies, respectively. Conclusion: This study found no significant differences in concordance among the tested assays for detecting anti-HPA and anti-HLA antibodies. These data suggest that no single method can detect all clinically important antibodies. Therefore, it is advisable that each laboratory develops customized protocols based on their expertise and employs complementary methods for comprehensive patient assessments.