Adi sod 111 f

After electrophoresis and protein transferring to nitrocellulose membranes, the membranes were blocked in Tris-buffered saline containing 0.1% Tween 20 and 3% BSA for 90 min at room temperature. Membranes were then incubated overnight at 4°C with primary antibodies, anti-glutathione Reductase (1:1000, ab137513, Abcam), anti-MnSOD (1:500, ADI-SOD-111-F, Enzo Life Sciences), anti-HSP70 (1:500, ADI-SPA-810-D, Enzo Life Sciences), anti-COX IV (1:500, ab33985, Abcam). After incubation with peroxidase-conjugated secondary antibodies, visualization was enhanced by chemiluminescence (GE Healthcare, anti-mouse IgG).

Each sample was loaded onto 4–20% Tris-glycine gels. After electrophoresis and transferring to nitrocellulose membranes, the membranes were blocked in Tris-buffered saline containing 0.1% Tween 20 and 0.2% I-block (Tropix, T2015) for 90 min at room temperature. Membranes were then incubated overnight at 4°C with following primary antibodies, anti-TOM40 (1:200, Santacruz, sc-11414), anti-glutathione reductase (1:500, Abcam, ab137513), anti-SOD2 (1:2000, Enzo Life Sciences, ADI-SOD-111-F). After incubation with peroxidase-conjugated secondary antibodies, visualization was enhanced by chemiluminescence (GE Healthcare, NA931- anti-mouse, or NA934- anti-rabbit, or NA935- anti-rat). Optical density was assessed using the NIH Image analysis software.