Nuclear factor κB luciferase assay

HEK293 cells were plated at 100 000 cells/well in a 24-well plate 24 h prior to transfection using transit-LT1. NFκB-luc (firefly luciferase containing proximal NFκB binding sites) and Renilla luciferase (pGL4 as a transfection control) were used at a 24 : 1 ratio for transfection, and 4 h after transfection, some cultures with were treated with 20 ng/mL tumour necrosis factor (TNF) (catalogue no. T0157; Milliport-Sigma, St Louis, MO, USA). At 24 h post-transfection, cells were harvested and luciferase activity assayed (Dual-Luciferase Reporter Assay System E1910; Promega, Madison, WI, USA).