GPIb-mediated activation of the GPIIb-IIIa complex is regulated by PECAM-1. (A) PECAM-1-deficient platelets (top row) show enhanced spreading on VWF compared with WT platelets (bottom row). Spreading was photographed and quantitated 30 minutes following addition of platelets to VWF-coated slides to accentuate the difference in spreading kinetics between wild-type and knock-out platelets. In the absence of botrocetin, platelets did not bind, and in the presence of RGD peptide, platelets bound but did not spread. The panels shown are representative fields from 2 independent, blinded experiments. Original magnification, ×100. (B) Digitized images of between 42 and 58 platelets per condition were acquired and analyzed using Metamorph software and showed a highly significant (2-tailed Student t test, ^*P < .001) difference in the size of spread PECAM-1-negative versus PECAM-1-positive platelets 30 minutes after addition to the VWF-coated surface. Error bars represent means ± SD. (C) Platelet agglutination-induced aggregation is enhanced in PECAM-1-deficient platelets (left panel), though the agglutination itself (VWF/botrocetin in the presence of RGD peptide [right panel]) is not significantly different.