Jam-B and Jam-C cytoplasmic tails are required for EC lumen and tube formation as well as Cdc42 activation in 3D collagen matrices. (A) ECs were infected with the indicated recombinant adenoviruses overnight and then were resuspended in 3D collagen matrices. After 24 hours, cultures were fixed, stained, and photographed. Arrows indicate EC lumens. Bar equals 50 μm. (B) EC lumens were quantitated by tracing lumen areas using Metamorph software. Data shown are normalized to the control sample and presents values of lumenal area per HPF ± SD. n = 10, P < .01. (C) Cultures were established as in panel A, and after 16 hours, detergent lysates were prepared from the 3D cultures and incubated with GST-PAK-PBD protein beads as described¹⁶  to assess the degree of Cdc42 activation. Starting material lysates as well as the eluates from the Pak beads were assessed by Western blot analysis using anti-Cdc42 antibodies.