Cell viability assay

The cell viability was evaluated by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) uptake method. Briefly, the 5-FU chemo-resistant cell lines, HT-29 and SW620, were seeded in a 96-well plate (1,000 cells per well) and exposed to different concentrations of 5-FU or 5-FU plus 5 μM morin, or 5-FU plus 3 μM MST-312 in triplicate for 24 h. Additionally, in another set of experiments, cells were co-treated with 5 μM morin and 3 μM MST-312 with different concentrations of 5-FU (0, 0.1, 1, 2, 3 and 4 μM) for 24 h to obtain the optimum dose for combination treatment. Cells were washed twice with PBS and subsequently, MTT solution (5 mg/ml) was added to each well and the plate was incubated for 4 h at 37°C. The 96-well plates were wrapped with aluminum foil and gently swirled for 15 min at room temperature. The absorbance of the cell suspension was measured at 570 nm. The data obtained were calculated and were represented as hundredth of survival relative to controls. This experiment was repeated 3 times independently, and statistical analysis was done to obtain the final values.