Mouse anti total oxphos antibody cocktail

Whole protein extracts of cholesterol- and mock-treated fibroblasts were prepared by lysing cells with RIPA buffer (Sigma Aldrich) containing a protease inhibitor cocktail (Roche). Proteins were separated by utilizing Bis-Tris Gels (Invitrogen) and transferred to PVDF membranes (Invitrogen). Membranes were blocked for 2 h at room temperature with 5% milk in PBS-T and incubated with primary antibodies at 4°C overnight. The primary antibodies and dilutions used are as follows: mouse anti-APP A4 1:1,000 (Merck-Millipore), mouse anti-GRP78 1:2,000 (BD Biosciences), rabbit anti-CASP3 1:1,000 (AB clonal), mouse anti-GAPDH 1:2,000 (Abcam), mouse anti-VDAC1 1:1,000 (Abcam), and mouse anti-total OXPHOS antibody cocktail 1:1,000 (Abcam). Protein signals were detected with a Super SignalTM West Pico PLUS (Thermo Scientific) kit according to the manufacturer’s protocol.