Anti-PECAM/conjugates cause PECAM-1 phosphorylation in endothelial cells. (A) HUVECs were incubated with either free anti-PECAM antibody or multivalent anti-PECAM/conjugates for 1 hour at 37°C. Cells were washed to remove nonbound conjugates, and tyrosine phosphorylation was determined from total cell lysates by Western blot using monoclonal HRP-labeled anti-pY, and goat polyclonal anti-PECAM followed by HRP-labeled anti–goat IgG. (B) Western blot densitometry. Dashed line indicates the level of PECAM-1 phosphorylation in HUVECs treated with anti-PECAM/conjugates at 4°C. Data are mean plus or minus SD (n = 3; *P < .05).