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. Author manuscript; available in PMC: 2014 Dec 19.
Published in final edited form as: J Biol Chem. 2005 Nov 22;281(4):2061–2070. doi: 10.1074/jbc.M507741200

Figure 6. Measurement of ROS production by CcO using modified DCF-DA method.

Figure 6

A shows the characteristics of the modified DCF-DA method for in vitro assay of peroxy radicals generated by liposome-reconstituted CcO enzyme. The rate of fluorescence emission by reactions with added DCF-DA alone (1 μm), and those with added reconstituted CcO (RV) from control heart mitochondria (100 μg), reduced cytochrome c (100 mM), partially purified cytosolic protein fraction rich in esterase activity (10 μg/ml), catalase (10 units/ml), and SOD (30 units/ml) are shown. B, the rate of fluorescence at the linear range (between 600 and 1400 s from A) was integrated to obtain the fluorescence units/min/mg protein. CcO from control and ischemic hearts reconstituted in lipid vesicles as described under “Materials and Methods” and Fig. 1 were used in the assay. Results in B represent average ± S.D. of four independent assays. Asterisks indicate significant difference (**, p < 0.01).