Rabbit anti caspase 1 antibody

Bone marrow-derived EPCs were lysed, centrifuged at 12, 000 g at 4 o C for 15 min, and the supernatant was obtained. We used BCA (Thermo, Rockford, USA) assay to determine the concentration of protein [28] . Samples containing equal amounts of protein were run on 10-15% SDS-PAGE. The proteins were electrotransferred to nitrocellulose filter membranes. The unbound sites in the protein membranes were incubated in PBS containing 5% non-fat dry milk for 4 h at 25 o C. After washing, the blots were incubated overnight at 4 o C with primary antibodies, and then incubated with secondary antibodies for 30 min at 25 o C in dark. Primary antibodies used included mouse anti-TXNIP antibody (1:500; International Co., Woburn, MA), rabbit anti-NLRP3 antibody (1:1000; ABCAm, Burlingame, CA), rabbit anti-caspase-1 antibody(1:200; Santa Cruz Biotechnology, Santa Cruz, CA), mouse anti-GAPDH (1:1000; Calbiochem, San Diego, CA) and mouse anti-actin (1:1000; Calbiochem, San Diego, CA). Secondary antibodies used included IRDye 800-conjugated anti-mouse antibody (1:5000; Rockland Immunochemical, Inc, Gilberts-ville, PA), and Alexa Fluor 680 goat anti-rabbit IgG antibody (1:5000; Invitrogen, Carlsbad, CA). The images were visualized using Odyssey infrared imaging system (Li-Cor Bioscience, Lincoln, NE), and the bands were quantified by Quantity One (Bio-Rad, Hercules, CA, USA).