Figure 6: Treatment with lapatinib and erlotinib generates reactive oxygen species (ROS) in ovarian cancer cells. (a) Heregulin treatment causes persistent elevation of ROS in ovarian cancer cells. Exponentially growing cells were seeded in triplicates in opaque flat bottom black-walled 96-well plates for 24 h. Following this, cells were either left untreated (UT) or treated with 1 nM heregulin for different time points as indicated. Following incubations, cells were loaded with DCFDA fluorescent stain for 45 min and assayed for ROS as described in Materials and Methods. (b) Lapatinib, erlotinib, and their combination cause ROS generation. Cells were seeded as in (a) and treated with either 1 nM HRG alone or with cotreatment of 5 μM lapatinib (LAP), erlotinib (ERLO), or their combination (COMB) for different time points as indicated, and ROS assay was repeated. For both (a) and (b), the fluorescence reading recorded from each well was normalised to total cell abundance within the same wells as described in Materials and Methods. Data shown are mean values ± S.D of triplicates, normalised to UT in (a) or HRG in (b) and expressed as fold change. Statistical significance was determined between treatment groups either by independent t-test or one-way ANOVA followed by post hoc Tukey’s test as appropriate and significance expressed according to the scale: (, , , and ).