Fig. 5. Effect of thrombin or TRAP on plasminogen binding to HUVECs. (A) Fluorescein-plasminogen (F-Plg) binding to HUVECs was evaluated following thrombin or TRAP treatment by incubating the cells with F-Plg for 30 minutes at 37°C. The cells were subsequently fixed, and DNA was stained with DAPI. Identical experiments were performed in which both media and F-Plg solutions contained either polyclonal anti-p11 IgG or non-immune rabbit IgG (rIgG). (B) Biotinylated plasminogen (B-Plg) binding to the surface of thrombin- or TRAP-treated HUVECs was evaluated by incubating cells with B-Plg for 30 minutes at 37°C. After washing unbound ligand, the cells were lysed and subjected to blot analysis using HRP-avidin. To ensure identical amounts of cell lysate were loaded in each lane, the blots were reprobed with anti-ß-actin antibody.