Differential Staining of Sister Chromatid in BrdU- Substituted Chromosomes after Incubation in Alkalified NaCl Solution

Chinese hamster strain cells cultured in the presence of BrdU (5-bromodeoxyuridine) for two or three successive generations are used. After hypotonic treatment and ethanol-acetic acid fixation, the slides are incubated at 80°C for 5 min in 2 M NaCl solution which had been alkalified by heating in a Coplin jar for several hours, and then stained with Giemsa. The procedure has resulted in dark staining of unifilarly BrdU-substituted (TB) chromatids and light staining of bifilarly substituted (BB) ones. The results of Feulgen staining are compatible with those of the Giemsa staining. This differential staining can reciprocally be converted by subsequent treatments with HCl followed by Giemsa staining to the type in which the BB-chromatids stain more intensely than the TB-chromatids.