RAW264.7 cells were treated with IFNβ (50 U/ml) for the indicated times (to induce expression of TRAFD1). 25 μg total cell lysate was loaded on a 10 % polyacrylamide gel and blotted overnight at 4 °C with anti-TRAFD1 (R3101) in 5 % milk at the indicated concentrations. Secondary HRP conjugated anti-Rabbit antibody was used at 1:10,000 dilution for 1 h at room temperature.