Department of Pediatrics, Division of Pediatric Neurology and Metabolism, Ghent University Hospital, De Pintelaan 185, 9000 Ghent, Belgium.
J Inherit Metab Dis. 2011 Jun;34(3):741-7. doi: 10.1007/s10545-011-9315-7. Epub 2011 Apr 12.
For more than a decade now blue native polyacrylamide gel electrophoresis (BN-PAGE) has been used for the study of the oxidative phosphorylation (OXPHOS) complexes. Catalytic activities of complexes I, II, IV and V can be assessed, after separation by gel electrophoresis, by incubation of the BN-PAGE gel in specific staining solutions. However, until now, a reliable staining method for testing ubiquinol cytochrome c oxidoreductase (complex III) activity by BN-PAGE gel techniques was not available. In addition, spectrophotometric methods currently in use for detection of complex III deficiency in patients are not very sensitive. Here, we describe a newly developed diagnostic method for visualization of complex III activity by direct in-gel evaluation of ubiquinol cytochrome oxidoreductase activity. We validated the method by reporting the results in six patients with previously characterised complex III defects.
十多年来，蓝色 native 聚丙烯酰胺凝胶电泳（BN-PAGE）一直被用于研究氧化磷酸化（OXPHOS）复合物。在凝胶电泳分离后，通过将 BN-PAGE 凝胶孵育在特定的染色溶液中，可以评估复合物 I、II、IV 和 V 的催化活性。然而，直到现在，BN-PAGE 凝胶技术检测泛醌细胞色素 c 氧化还原酶（复合物 III）活性的可靠染色方法还不可用。此外，目前用于检测患者复合物 III 缺乏的分光光度法不是很敏感。在这里，我们描述了一种新开发的诊断方法，通过直接在凝胶中评估泛醇细胞色素 c 氧化还原酶活性来可视化复合物 III 活性。我们通过报告六名先前特征为复合物 III 缺陷的患者的结果验证了该方法。
