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. 2017 Mar 10;12:462–468. doi: 10.1016/j.redox.2017.03.002

Fig. 2.

Fig. 2.

Generation of superoxide free radical anion (O2•–) and H2O2 by eGFP in the presence of NADH. (a) Using eGFP expressed by E. coli, rates of NADH consumption and O2•– formation were determined in samples containing 15 μM eGFP, 150 μM NADH and 20 μM cytochrome c. The effects of catalase (1 kU/mL) and SOD (250 U/mL) were also studied. O2•– formation was determined by the cytochrome c reduction measured at 550 nm (ε550 nm=21,000 M−1 cm−1[34]). (b) Rates of NADH consumption and O2•– formation were studied in samples containing 15 μM GFP, 20 μM cytochrome c and different concentrations of NADH. (c) H2O2 production was determined in samples containing 25 μM GFP and 250 μM NADH. Every 5–6 min, an aliquot was withdrawn and H2O2 measured. Samples to which catalase (1 kU/mL) had been added were assayed in parallel. (d) Using eGFP expressed by the mammalian cell line HEK 293, the rate of O2•– formation was determined in samples containing 15 μM eGFP, 150 μM NADH and 20 μM cytochrome c, such as described in (a). The effect of SOD (250 U/mL) was also studied. * is used to show statistically significant differences between samples and controls.