[3H]thymidine incorporation assay

To examine the effect of the compounds on cell proliferation, MCF-7 cells were seeded (2×10⁶) in 6-well plates and cultured as described. Cell cultures were incubated with varying concentrations of the tested compounds and 0.5 µCi [³H]thymidine for 24 h at 37°C. The cells were harvested by trypsinization and washed several times in cold PBS (10 min/1.500 g) until the dpm in the washes were similar to the reagent control. Radioactivity was determined by liquid scintillation counting. [³H]thymidine uptake is expressed as dpm/well (9).