Characterization of platelet-specific alloantigens by immunoblotting: localization of Zw and Bak antigens.

The glycoprotein localization of the platelet-specific antigens Zwa, Zwb and Baka and their presence on tryptic fragments of glycoproteins was studied by immunoblotting. Human platelets were solubilized and pre-cleared from platelet-associated IgG. The glycoproteins were separated on SDS polyacrylamide gels, transferred to nitrocellulose and incubated with platelet antibodies, followed by 125I-radiolabelled anti-human Ig antibodies. Glycoprotein IIb/IIIa were isolated from platelet lysates by immuno-affinity chromatography. These proteins were subjected to trypsin digestion, and then used for the immunoblot procedure with platelet antibodies. A glycoprotein specifically reacting with either anti-Zwa or anti-Zwb was found, with an apparent molecular weight of 88 kDa. This protein co-migrated, and was probably identical with, glycoprotein IIIa. After trypsin digestion the smallest fragment, reactive with IgG anti-Zwa or IgM anti-Zwb, had a molecular weight of approximately 23 kDa. IgG anti-Baka and anti-Leka antibodies reacted with a protein of 130 kDa from platelets of Bak(a+) donors. This protein was identified as glycoprotein IIb.