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doi: 10.1172/JCI96113 204 TangHLiangYAndersRATaubeJMQiuXMulgaonkarAet al

Cell viability and ROS and glutathione intracellular content of cells cultured in liquid medium were assessed by flow cytometry analysis of cells stained for 30 min in the dark with 0.025 M calcein-AM (ThermoFisher Scientific, C3099) at room temperature, 50 M Dichlorodihydrofluorescein-diacetate (DCFDA, ThermoFisher Scientific, D399) at 37 C, and 10 M monochlorobimane (ThermoFisher Scientific, M1381MP) at 37 C, respectively

Subsequently, all samples, including the diamide-treated control and test samples, containing approximately 1 million cells each, were incubated with 250 L of 80 M H2DCF-DA prepared in DMSO dye in the same buffer for 20-30 minutes at 30 degree C in the dark

It was more effectively activated by elevated levels of GSH in H1299 lung cancer cells than that in normal lung fibroblasts WI38 cells, releasing reactive alkoxyamines to capture AP sites, blocking APE1 repair, and leading to lung cancer cell death in vitro