Binding of GST-Grb2 but not GST to WASP. Platelets were stimulated in nonaggregating conditions and lysed as described for Fig1. Lysates before (−) and 5 minutes after (+) collagen stimulation were equally divided into four aliquots. The two samples from each of the lysates were incubated either with agarose-bound GST-Grb2 (10 μg, lanes 1 and 2) or GST (10 μg, lanes 3 and 4) on ice for 3 hours. After three washes in phosphate-buffered saline (150 mmol/L, pH 7.4) containing orthovanadate (200 μmol/L), the precipitates were incubated in SDS sample buffer. The SDS samples were subjected to Western blotting to detect WASP by polyclonal anti-WASP antiserum 503.