Binding of biotinylated alboaggregin A to platelet lysate.

Platelet lysate was separated by SDS-PAGE (7%-17% acrylamide gradient) and transferred to PVDF membranes. Proteins were incubated with (A) anti-GPVI Ab followed by antirabbit IgG–coupled alkaline phosphatase, (B) biotinylated alboaggregin A and avidin-coupled alkaline phosphatase, or (C) avidin-coupled alkaline phosphatase and detected with BCIP and NBT. (D) Proteins from platelet lysate immunoprecipitated with rabbit preimmune serum anti-GPVI (lane 1), rabbit anti-GPV Ab (lane 2), rabbit anti-GPVI Ab (lane 3), or isolated GPVI purified on a convulxin column (lane 4) was separated by SDS-PAGE (7%-17% acrylamide gradient) and transferred to PVDF membranes. The proteins were incubated with biotinylated alboaggregin A before detection with avidin-coupled alkaline phosphatase, BCIP, and NBT.