Anti sdha

25 μg of islet or mitochondrial protein lysates were separated on a Criterion-TGX stain-free gel (Bio-Rad). Immunoreactive proteins were visualised on a Fuji-Film LAS4000 and Gel-Doc Ez-Imager (Bio-Rad). Primary antibodies; anti-RCAN1 (1:200, Sigma Aldrich), anti-TOM20 (1:1000, Jackson Laboratories), anti-β-actin (1:200, Sigma Aldrich), anti-NDUFA9 (1:1000, Sigma Aldrich), anti-SDHA (1:2000, Sigma Aldrich), anti-CORE1 (1:1200, Sigma Aldrich) and anti-Opa1 (1:1500, Sigma Aldrich). Secondary antibodies; donkey anti-rabbit horse-radish peroxidase (HRP) (1:100, Life Technologies) and donkey anti-mouse HRP (1:100, Life Technologies).

Blue native polyacrylamide gel electrophoresis (BN-PAGE) was conducted as mentioned before [27 (link)]. For mitochondrial supercomplexes, 1% digitonin (Sigma-Aldrich) was used to dissolve the membrane protein from patient and controls, then 3.5–16% gradients polyacrylamide gel to separate the supercomplexes. For mitochondrial complexes, 1% dodecyl maltoside (DDM, Sigma-Aldrich) was used and then 3–11% gradient polyacrylamide gel to separate the complexes. The proteins were transferred on the PVDF membrane (Bio-Rad, USA), blocked with 5% no-fatty acid milk (Mengniu, China), incubated with the first and second antibodies, and then detected the protein signals using. The following antibodies were used: anti-Grim19 (1:1000, Sigma-Aldrich), anti-SDHA (1:3000, Sigma-Aldrich), anti-UQCRC2 (1:2000, Sigma-Aldrich), anti-MT-COI (1:2000, Sigma-Aldrich), anti-ATP5A (1:3000, Sigma-Aldrich), anti-NDUFB6 (1:2000, Sigma-Aldrich), anti-NDUFS3 (1:2000, Sigma-Aldrich), anti-TOM70 (1:2000, Proteintech, China), anti-mouse IgG HRP linked (1:2000, Cell Signaling Technology), anti-rabbit IgG HRP linked (1:2000, Cell Signaling Technology), anti-mouse IgG, AP (1:2000, Cell Signaling Technology).