Succinate dehydrogenase (SDH) commonly is assayed as a marker enzyme for mitochondrial activity. The literature presents numerous conditions for conducting this assay due to the fact that, it has been difficult to get sufficient reduction of the acceptor dye, 2,3,5-triphenyl-2H-tetrazolium chloride (TTC). This study was undertaken to optimize the SDH-catalyzed reduction of TTC dye by evaluation of a greater range of molor ratios of TTC to succinate and by further evaluation of additives reported as beneficial. Improvement in enzyme specific activity was achieved by liver perfusion via the left cardiac ventricle with homogenizing solution. Increase in TTC from 1 to 10 mM and further increase to 20 mM resulted in major improvement in color production. The greatest improvement in apparent activity was achieved by addition of 1 mM phenozine methosulfate, a hydrogen transfer mediator. Use of CaCI₂. EDTA, Triton X-100, NaN₃ and KCN was not beneficial. The above modifications of the SDH assay resulted in greater sensitivity, the conduct of a greater number of assays with less tissue and the sacrifice of fewer animals.
Shaw, Collie B.; Chronister, Tara L.; and Peck, John D.
"Optimal Conditions for Kinetic Study of Succinate Dehydrogenase in Rat Liver,"
Journal of the Arkansas Academy of Science: Vol. 40
, Article 21.
Available at: http://scholarworks.uark.edu/jaas/vol40/iss1/21