Nih3t3 mefs

HEK293T/17 cells and NIH3T3 MEFs (all ATCC) maintained in DMEM containing 10% fetal bovine serum, 4 mM L-glutamine, 100 U/mL penicillin and streptomycin (all Gibco) under 5% CO2, atmospheric oxygen, at 37°C in a humidified incubator.

NIH 3T3 MEFs were purchased from ATCC. The 3T8 Jurkat T cell line has been described elsewhere (33 (link)) and was previously mutagenized to generate the NEMO^KO cell line 8321 (33 (link)). These 8321 cells were transfected with plasmid encoding full-length NEMO to generate 8321^WT cells (30 (link)). All fibroblasts and Plat-E cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM, Invitrogen) supplemented with 10% fetal bovine serum (FBS, Invitrogen), 2 mM L-glutamine, penicillin (50 U/ml), and streptomycin (50 U/ml). Jurkat T cells were cultured in RPMI medium (Invitrogen) supplemented with 10% FBS, 2 mM L-glutamine, penicillin (50 U/ml), and streptomycin (50 U/ml). Cells were passaged with 0.25% trypsin (Invitrogen). Unless otherwise noted, cells were stimulated with TNF-α (10 ng/ml), LIGHT (100 ng/ml), or anti-LTβR antibody (300 ng/ml) when they reached 80% confluence.