Mouse monoclonal igg antibody anti p65

Samples taken from either cells or tissues of mice after experimental treatment were lysed with RIPA buffer (30 mM Tris-HCl, 150 mM NaCl, 2 mM EDTA, 1% Triton X-100, 10% glycerol, and complete cocktail (Life technologies, Grand Island, NY) and phosphatases (Sigma, St. Louis, MO). Lysates were centrifuged at 14000×g for 15 min, the supernatants were collected and the concentration was quantitated. The samples were boiled for 10 min, and equal amount was applied to 12% SDS-polyacrylamide minigels and electrophoresed. The proteins in the gel were then transferred to nitrocellulose filter membranes (Thermofisher, Rockford, IL). Horseradish peroxidase (HRP)-linked secondary antibody (Rockland, Gilbertsville, PA) and X-ray film (Kodak) were used for exposure ^{47 (link),48 (link)}. Rabbit polyclonal IgG antibody anti-GAPDH, c-Myb, MyD88, TLR4, PKA, pPKA (Thr 198), CREB-1, pCREB-1 (Ser 133), pp65 (Ser 536), TGF-β1, goat polyclonal IgG antibody anti-IL-4, and mouse monoclonal IgG antibody anti-p65 were bought from Santa Cruz Biotechnology (Santa Cruz, CA). Rabbit polyclonal IgG antibody anti-TRIF were bought form Abcam (Cambridge, MA).