Retinas were homogenized in RIPA buffer containing 1% Triton X-100, 5% SDS, 5% deoxycholic acid, 0.5 M Tris HCl, pH 7.5, 10% glycerol, 1 mM EDTA, 1 mM phenylmethylsulfonyl fluoride (PMSF), 5 × 10−12 μg/ml aprotinin, 1 × 10−12 μg/ml leupeptin, 1 × 10−12 μg/ml pepstatin, 200 mM sodium orthovanadate, and 200 mM sodium fluoride. Tissue extracts were incubated on ice for 10 minutes and centrifuged at 10,000g for 25 minutes at 4°C. Total protein in retinal extracts was measured using a standard BCA assay (Pierce, Rockford, IL), and the protein concentration was determined by the Lowry method (Bio-Rad Life Science, Mississauga, ON, Canada). Retinal extracts were resuspended in 5× sample buffer (60 mM Tris HCl, pH 7.4, 25% glycerol, 2% SDS, 14.4 mM 2-mercaptoethanol, 0.1% bromophenol blue), boiled for 5 minutes, and resolved on a 12% SDS-PAGE gel. Proteins were transferred onto a nitrocellulose membrane (Hybond-C; GE Healthcare, Little Chalfont, Buckinghamshire, UK), and blots were stained (Ponceau S; Sigma-Aldrich) to visualize the protein bands and to ensure equal protein loading and uniform transfer. The blots were then washed with Tween-Tris-buffered saline solution and were blocked with 5% nonfat dry milk in TBST buffer (20 mM Tris HCl, pH 7.6, 137 mM NaCl, and 0.1% Tween 20) for 45 minutes. The blots were probed with the primary antibody against IL-1β (1:1000 dilution; Sigma-Aldrich) for 24 hours, followed by the horseradish-peroxidase (HRP)-conjugated goat anti-rabbit secondary antibody (1:10,000 dilution). Bound antibodies were detected using an enhanced chemiluminescence system (ECL; GE Healthcare) and x-ray film. Signal intensity was measured (ImageMasterRVDS; GE Healthcare), and the optical densities (mean ± SD) for each sample were obtained from three measurements on three separate blots.