Endothelial cells assemble prominent focal adhesions on αC(FXIII) and αC(tTG) oligomers but not on αC monomers. HUVECs in serum-free DMEM were plated on 20 μg/mL αC monomers (A-B), αC(FXIII) oligomers (C-D), or αC(tTG) oligomers (E-F) for 2 hours. Paraformaldehyde-fixed, Triton X-100–permeabilized cells were double stained with anti-β3 integrin mAb 25E11 and polyclonal antiphosphotyrosine antibodies, followed by rhodamine-labeled anti–mouse and fluorescein-conjugated anti–rabbit IgG. A clear peripheral staining for αVβ3 integrin and phosphotyrosine was observed at lower magnification in HUVECs on αC oligomers (C,E) but not on monomeric αC domains (A). At higher magnification, well-developed focal contacts containing αVβ3 integrin and phosphotyrosine were detected in HUVECs on αC oligomers (D,F), whereas no distinct focal adhesions were formed on αC monomers (B). Bars indicate 20 μm. Arrowheads mark colocalization of αVβ3 integrin and phosphotyrosine in the peripheral focal contacts of HUVECs on αC oligomers.