Comparison of synchronized Chinese hamster ovary cells obtained by mitotic shake-off, hydroxyurea, aphidicolin, or...

Synchronized cell populations are necessary to study many aspects of cell biology. We have developed a method to obtain highly synchronized Chinese hamster ovary cell populations in S phase or G2 phase by utilizing mitotic selection followed by incubation with either hydroxyurea, aphidicolin, or methotrexate for 12 h. Flow cytometry analysis shows that the coefficient of variation in the spread of the cell population in S phase is as low as 6%. Drug toxicity studies compare the effects of the various drugs on G1 and S phase cells. The use of aphidicolin or hydroxyurea results in the most highly synchronized cell populations, but methotrexate yields inadequate synchronization. These results demonstrate that both aphidicolin and hydroxyurea are useful drugs for obtaining highly synchronized cell populations after an initial synchrony in mitosis. Aphidicolin is perhaps the best choice because of less toxicity to S phase cells when used in low concentrations.