CaSki cells display higher levels of ROS than do SiHa cells (a), the cell viability of CaSki cells decreases more than in SiHa cells after treatment with tBHP (b). Pretreatment with NAC decreases ROS in CaSki and SiHa cells (c) and pretreatment with NAC protects cells from death induced by treatment with tBHP (d). (a) 105 cells of each line were treated with 10 μM DCFDA or 10 μM DHE in media and then incubated in the dark at 37°C for 30 minutes. The cells were washed and, after trypsinization, were resuspended in 1x PBS and analysed by flow cytometry. A total of 10 000 events were measured per sample. (b) SiHa and CaSki cells (1.5 × 104 cells per well) were seeded into a 96-well plate, allowed to incubate overnight, and then treated with the indicated concentrations of tBHP for 20 h. Viability was measured by crystal violet. The viability of untreated cells was set at 100%. Each measurement was done in triplicate and error bars indicate the standard deviations of the means. (c and d) Pretreatments with NAC were begun 24 h prior to treatment with tBHP (d). (c) The measurement of ROS levels was performed as described in (a) and (d) the measurements of cell viability were performed as described in (b).