On the association of the platelet-specific alloantigen, Pen(a), with glycoprotein IIIa. Evidence for heterogeneity of glycoprotein IIIa

Abstract

Neonatal alloimmune thrombocytopenic purpura associated with a new platelet-specific alloantigen Pen(a) has been reported. We now provide direct evidence that the Pen(a) determinant is associated with glycoprotein (GP) IIIa, but that it is distinct from epitopes that define the Pl(A) system. By ELISA wherein monoclonal antibodies specific for GPIIb (Tab) and specific for GPIIIa (AP3) were used to capture and hold antigens from a platelet lysate prepared under conditions that generate free GPIIb and GPIIIa, anti-Pen(a) reacted with GPIIIa held by AP3 but not with GPIIb held by Tab. In an alternative ELISA where purified GPIIIa from both Pl(A1)-positive and Pl(A1)-negative platelets were used individually as antigen, anti-Pen(a) reacted with both allelic forms of GPIIIa. By radioimmunoprecipitation, anti-Pen(a) precipitated a single surface-labeled membrane protein with electrophoretic characteristics in sodium dodecyl sulfate-polyacrylamide gels, under nonreduced or reduced conditions, identical to those of GPIIIa. By fluorocytometry, platelets from several donors, regardless of Pl(A) phenotype, bound an amount of anti-Pen(a) roughly equivalent to one-half that amount of anti-Pl(A1) bound by Pl(A1) homozygous (A1/A1) platelets and roughly equal to that amount of anti-Pl(A1) bound by Pl(A1) heterozygous (A1/A2) platelets. Using platelets from donors typed homozygous for Pl(A1) and Pen(a) in a quantitative indirect binding assay, 14-24,000 molecules of anti-Pen(a) and 41-51,000 molecules of anti-Pl(A1) were bound per platelet at saturation. Anti-Pen(a) completely inhibited ADP-induced aggregation of Pen(a)-positive platelets, regardless of Pl(A) phenotype. These results indicate that the Pen(a) determinant is associated with GPIIIa but distinct from Pl(A).