Caspase-8/FLICE is upregulated and activated by IFNγ. In two separate experiments, day-6 cells were cultured in medium with or without 500 U/mL of IFNγ for 24 to 96 hours. Cell protein lysates were prepared and immunoblot analysis was performed with anti–caspase-8 antibodies. Each lane contained 150 μg of total protein. The top panel, using antibody to Mch5 p10 and a 3-minute exposure, shows that pro–caspase-8 increased after incubation with IFNγ for 24 hours. The bottom panel shows the result with antibody to Mch5 p20 and 15 minutes of exposure. Activation products of caspase-8, p31, and p20 are clearly observed in the IFNγ-treated cells by 24 hours.