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The diagnosis of mitochondrial encephalomyopathies caused by complex I (C-I) deficiency relies mainly on the spectrophotometric C-I assay. Considered difficult, this assay lacks reliability and has high nonspecific activity. We studied the key factors of this assay in cultured cells (cybrid and fibroblast): ubiquinone analogues, rotenone inhibition to determine specific activity, and mode of permeabilization of mitochondrial membranes. We showed that ubiquinone 1 allows a more stable and reliable assay, better results were obtained with rotenone inhibition done in the same assay, and mitochondrial permeability was improved just by freeze/thawing the sample prior to the assay. Copyright © 2012 Elsevier Inc. All rights reserved.


Kátia Klug Oliveira, Beatriz Hitomi Kiyomoto, Andresa De Santi Rodrigues, Célia Harumi Tengan. Complex I spectrophotometric assay in cultured cells: detailed analysis of key factors. Analytical biochemistry. 2013 Apr 1;435(1):57-9

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PMID: 23246343

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