NF-κB activation by free or fibrinogen-bound IL-1β Endothelial cells were plated on gelatin-coated wells in McCoy 5A medium supplemented with 20% FBS, 50 μg/mL endothelial cell growth supplement (ECGS), and 100 μg/mL heparin and grown to confluence. The cells were then washed twice with McCoy medium and incubated with 2 ng/mL of IL-1β in the presence or absence of 1 μg/mL fibrinogen for 2 hours. Nuclear protein extracts were then prepared, and EMSA was performed using a double-stranded ³²P-labeled consensus NF-κB primer. LPS (1 μg/mL) was used as a positive control. Lanes are as follows: 1, LPS; 2, medium alone; 3, fibrinogen; 4, IL-1β; and 5, fibrinogen plus IL-1β.