DEVELOPMENT AND EVALUATION OF A SPECTROPHOTOMETRIC ASSAY FOR COMPLEX III IN ISOLATED-MITOCHONDRIA, TISSUES AND FIBROBLASTS FROM RATS AND HUMANS

A spectrophotometric method to assay the activity of complex III in is olated mitochondria, tissues and fibroblasts from patients and rats ha s been developed and validated, Decylubiquinol was shown to be a suita ble substrate with a saturating concentration between 100 and 200 mu m ol/l. The optimal pH was found to range from 7.4 to 8.0. The enzyme re action was linear for incubations containing up to 15 mu g/ml mitochon drial protein, 250 mu g/ml liver tissue, 375 mu g/ml skeletal muscle o r 100 mu g/ml fibroblast protein. Intraday and interday variability of the assay for different enzyme sources was below 15% and 10%, respect ively. Assessment of complex III activity in liver and fibroblasts fro m patients with signs of mitochondrial dysfunction revealed the useful ness of the newly developed assay in the diagnosis of complex III defi ciency,