Recombinant human PECAM-1–immunoglobulin chimera can induce tyrosine phosphorylation of PECAM-1.

(A) IV.3-treated washed platelets were stirred in the presence of 40 μg/mL recombinant human PECAM-1–immunoglobulin chimera for 0 to 6 minutes. Platelet lysates were precleared with Protein G–Sepharose beads, PECAM-1 immunoprecipitated, separated by SDS-PAGE, and immunoblotted for anti-phosphotyrosine content using an HRP-conjugated 4G10 anti-phosphotyrosine antibody. The presence of PECAM-1 antigen was confirmed by reprobing with polyclonal anti–PECAM-1 antibody SEW16. (B) PECAM-1 was immunoprecipitated from human platelets under resting conditions or after stimulation with CRP (2 μg/mL) (0-6 minutes) and 2 μg/mL CRP + 40 μg/mL recombinant human PECAM-1–immunoglobulin chimera (0-6 minutes). Proteins were separated on SDS-PAGE and immunoblotted for anti-phosphotyrosine content using an HRP-conjugated 4G10 anti-phosphotyrosine antibody. The presence of PECAM-1 antigen was confirmed by reprobing with polyclonal anti-PECAM-1 antibody SEW16.