PAK^TM-Lx: A New Luminex*^®-Based Assay to Detect and Identify Antibodies to Human Platelet Antigens (HPA) in Patient Serum.

Abstract 4187

Platelets express a variety of glycoproteins (GPIIb-IIIa, GPIa-IIa, GPIb/IX, GPIV, CD109 and Class I HLA). Human Platelet Antigens (HPA) represent polymorphisms of these platelet surface glycoproteins. While these polymorphisms do not alter the function of the proteins, they can become target of the immune system in mismatched individuals. Alloantibodies against HPA are involved in the pathogenesis of Neonatal Allo-Immune Thrombocytopenia (NAIT), Post-Transfusion Purpura (PTP) and refractoriness to random donor platelets. In addition to generating anti-HPA antibodies, these patients also develop antibodies to Class I Human Leukocyte Antigen (HLA). Most reference laboratories now use a combination of a sensitive screening assay such as the Platelet Immunofluorescence Test (PIFT) along with a solid phase assay for antibody identication, usually either the ELISA and Monoclonal Antibody Immobilization of Platelet Antigens (MAIPA) as well as genotyping techniques for at least HPAs 1, 2, 3, and 5. Currently, MAIPA is considered the gold standard in detection of antibodies to HPA. However, all existing methods have significant limitations in terms of sample requirements, time and labor involved. These factors are a major impediment as to how many specificities can be resolved in a rapid, efficient and reproducible manner. Development of microbead-based assays like the xMAP (Luminex^®) has provided a technology that can resolve these questions. GTI Diagnostics^® and Gen-Probe have developed PAK^TM -Lx, a bead-based assay for the Luminex^® platform to detect and determine the specificity of anti-HPA antibodies. PAK^TM -Lx is capable of resolving the specificities of antibodies to HPA 1, 2, 3, 4 and 5 in addition to detecting antibodies against GPIV and HLA Class-I antigens. Furthermore, it is capable of detecting antibodies to multiple HPA and HLA with only 10 μl of sample. With a total time of less than two hours from addition of samples to end results, PAK^TM -Lx is sensitive, rapid and simple to perform. Focused, flexible multiplexing in the range of 1 to 100 analytes provided by Luminex^® technology meets the needs to expand the panel to include more HPA combinations, thereby reducing sample requirements, labor and costs.