An improved spectrophotometric method for a more specific and accurate assay of mitochondrial complex III activity

BackgroundThe determination of the activity of complex III in tissue samples provides critical evidence in the diagnosis of mitochondrial disorders and diseases associated with mitochondrial dysfunction. However, great variations have been seen in the literature due to the use of different assays.MethodsReaction conditions of an improved spectrophotometric method exhibiting higher specificity for complex III activity than the methods currently used, were studied.ResultsOptimum conditions, using bovine serum albumin at 0.01% and Tween-20 at 0.05%, were defined. The present method possesses more antimycin A-sensitive complex III activity, compared to previous methods. Thus, this improved method is sensitive and suitable for assaying complex III in both crude tissue homogenate and isolated mitochondria of liver, heart, skeletal muscle and brain.ConclusionsThis spectrophotometric assay is sensitive, and specific for complex III activity because of the negligible blank rate and high antimycin A-sensitive activity. The low concentration of bovine serum albumin, and the use of inexpensive detergent Tween-20 make this improved method more robust for its use in a clinical laboratory setting.