Cybrids and OXPHOS Biochemical Assays

Fibroblast cells were grown in DMEM medium supplemented with glutamax (446 mg/l), 10% fetal calf serum, 50μg/ml uridine and 1mM sodium pyruvate under standard conditions. Cells were fused with 143B rhoo cells as previously described [63 (link)]. Several clones were isolated and the resulting cybrid cells were subsequently expanded. Biochemical assays were performed on isolated mitochondria and/or permeabilized cells. Western blot analysis of a patient’s muscle biopsy sample and Blue Native gel analysis (BNG) of a patient’s fibroblast were performed as previously described [63 (link)]. We used “MitoProfile” antibody mix (Total OXPHOS human WB antibody cocktail, Abcam) for muscle lysates. For fibroblast lysates we used antibody GRIM19 (Abcam), which corresponds to the mitochondrial complex I subunit NDUFA13.