Iκbα rabbit polyclonal antibody

Following stimulation with LPS (Enzo Life Sciences), Cells were washed with cold PBS, gently scraped, and resuspended in lysis buffer (1.0% NP-40, 150 mM sodium chloride, 50 mM Tris, pH8.0) containing proteinase inhibitor cocktail (Sigma-Aldrich) and phosphatase inhibitor cocktail 2 and 3 (Sigma-Aldrich). Protein samples were mixed with laemmli sample buffer (Bio-Rad), resolved by 12% SDS-PAGE gel, and transferred to nitrocellulose membranes (Millipore). The membrane was probed by phospho-NF-κB p65 (Ser536) rabbit monoclonal antibody (Cell signaling, #3033; 1:1000 dilution), NF-κB p65 rabbit polyclonal antibody (Santa Cruz, #sc-372; 1:1000 dilution), Phospho-IRF3 (Ser396) rabbit monoclonal antibody (Cell signaling, #4947; 1:1000 dilution), IRF3 rabbit monoclonal antibody (Cell signaling, #4302; 1:1000 dilution), IκBα rabbit polyclonal antibody (Santa Cruz, #sc-371; 1:1000 dilution), and α-actin goat polyclonal antibody (Santa Cruz, #sc-1616; 1:1000 dilution).

All reagents were purchased from Sigma-Aldrich (St. Louis, MO, USA), if not specified. RPMI1640, MEM, fetal bovine serum (FBS), penicillin/streptomycin, and trizol reagent were purchased from Invitrogen Life Technologies (Carlsbad, CA, USA). Antibody of Lamin B, I-κB-α rabbit polyclonal antibody, β-actin mouse monoclonal antibody, NF-κB p65 rabbit polyclonal antibody, phospho-ERK antibody, MMP-7 goat polyclonal antibody, p67phox goat polyclonal antibody, NOX1 rabbit polyclonal antibody, NOX2 goat polyclonal antibody, celecoxib were obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Phospho-p38 MAPK antibody, ERK antibody, p38 MAPK antibody, phospho-AMPK-α, AMPK-α, phospho-JNK, JNK, C-Jun, phospho I-κB, and PD98059 were purchased from Cell Signaling Technology, Inc. (Boston, MA, USA). Matrigel was obtained from BD Biosciences (Bedford, MA, USA). Trypsin/EDTA was purchased from Clonetics, Inc. (Walkersville, MD, USA). SR11302 was purchased from Tocris Bioscience (Tocris House, Bristol, BS110QL, UK). D942 was purchased from Calbiochem (10394, Pacific Center Ct, CA, USA).