A PLATELET MONOCLONAL-ANTIBODY INHIBITION ASSAY FOR DETECTION OF GLYCOPROTEIN IIB/IIIA-RELATED PLATELET ALLOANTIBODIES

Post-transfusion purpura (PTP) and neonatal alloimmune thrombocytopenia (NAT) result from formation of alloantibodies to platelet membrane glycoprotein-associated antigens. The detection and identification of platelet-specific alloantibodies in patient sera is often complicated by the presence of co-existing HLA antibodies and/or more than one platelet specificity in the same serum. We describe a solid phase assay that specifically detects antibodies to platelet membrane associated alloantigens by measuring the ability of patient antisera to inhibit the binding of glycoprotein GPIIb or GPIIIa monoclonal antibodies to intact platelets. When tested in the GPIIIa assay against a panel of random platelet donors, the reactivities of two known PL(A1) antisera that also contained different HLA antibodies were highly correlated (r = 0.99) and allowed PL(A) phenotyping of the population. A standard direct binding platelet ELISA, on the other hand, was unable to accurately PL(A) phenotype the same population. The reactivities of two known Bak(a) antisera (one containing additional anti-PL(A2) and the other anti-Br-b specificities) were highly correlated (r = 0.95) in the GPIIb assay, and Bak phenotype determination was similarly accomplished for a random platelet panel. Furthermore, a comparison of platelet phenotype results (using the monoclonal inhibition assay) and genotype results (using DNA analysis) for the PL(A) and Bak systems showed a concordance of 98% for 146 alleles tested. In conclusion, the platelet monoclonal antibody inhibition assay: (1) allows determination of platelet-specific alloantibodies in the presence of contaminating HLA antibodies and/or in sera containing multiple platelet alloantibodies; (2) allows accurate platelet phenotyping for the GPIIIa-associated PL(A) and GPIIb-associated Bak antigen systems; and (3) may be applicable to the detection of other known or even novel platelet glycoprotein-associated antigens.