Fig. 6.

Effects of BrdU treatment on cell viability. a The bar graph shows colony formation assays of wt JM8.N4 ESCs to test effects of BrdU and BrdU/dCTP combinations at indicated concentrations. BrdU/dCTP concentrations are shown in micromolars. Cells were plated at equal densities on 6-well dishes (day 0), and after 1 day (day 1), were exposed to BrdU, dCTP, or BrdU/dCTP. Treatment time was 7 h, except for 40-μM BrdU (16 h). dCTP exposure was kept for another 24 h and was subsequently reduced in 50 % increments for two more days. On day 2, cells were trypsinized and split either 1:65,000 or 1:100,000, and colonies were counted after 1 week. Colony numbers were normalized to untreated control samples. Averages of three experiments are shown including error bars for standard deviations, except for one sample (1.25/0 μM BrdU/dCTP, indicated by an asterisk) where the experiments were only carried out twice. b In one experiment, cells were split into 6-cm plates on day 2, and 7 days later colonies were stained for alkaline phosphatase (AP) activity. All colonies were AP positive