BIOTIN-AVIDIN IMMOBILIZATION OF PLATELET GLYCOPROTEINS (BAIPG) - A NEW CAPTURE ASSAY FOR THE DETECTION OF ANTIPLATETLET ANTIBODIES

Several 'capture' assays are currently employed to identify specific p latelet antibodies, but all require the use of murine monoclonal antib odies (MoAbs) against the antigen of interest. We have developed a new antigen capture assay for the detection of platelet reactive antibodi es, based on platelet surface sialoglycoprotein labelling with biotin hydrazide, and a following immobilization of the biotinylated platelet proteins to microtiter wells that had been coated with streptavidin. The resulting solid phase can then be used in a simple ELISA to detect serum and platelet associated antibodies. We describe here two versio ns of this biotin-avidin immobilization of platelet glycoproteins (BAI PG) assay. In BAIPG assay type I, the test sera are directly incubated in microtiter wells previously coated with streptavidin plus biotinyl ated platelet proteins. The BAIPG type II procedure involves the incub ation of sera with biotinylated platelets before platelet solubilizati on, and, after platelet lysis, the immobilization of the immune comple xes to streptavidin-coated wells. In both cases, the bound antibodies are determined by alkaline phosphatase conjugated anti-human IgG. Usin g BAIPG type I, positive results were obtained in 7/33 patients with i diopathic thrombocytopenic purpura (ITP), 1/10 patients with secondary immune thrombocytopenia (SIT) and 4/17 with non-immune thrombocytopen ia (NIT). The BAIPG type II test was positive in 13 out of 33 patients with ITP, in six out of ten patients with SIT, and in three out of th e 17 patients with NIT. A comparison between BAIPG and monoclonal anti body immobilization of platelet antigens (MAIPA) assays showed a high degree of correlation between the two methods. These results suggest t hat the BAIPG assay is a valuable new tool for the detection of anti-p latelet antibodies.