Tritiated thymidine continuous labelling

Detailed analysis using several different methods (Pulse and continuous label with tritiated thymidine, DNA fluorescence per cell using the fluorescent Feulgen assay and flow microfluorography (Van Dilla et al., 1969), and DNA specific fluorescence using the diaminobenzoic acid assay (Kissane and Robins, 1958) reveals that the DNA content of a cell increases in a saltatory fashion, and that the early portion of S-phase is a period of low net DNA synthesis which may be mistaken for G1 if insensitive methods of measurement are used (Klevecz et al., 1975). 

Fig. 10.8. Continuous labelling method. Human skin epithelial cell cultures were exposed to tritiated thymidine (2.5 /iCi/ml 0.06 Ci/mol) continuously for 25 h during which time cells were harvested and scored for the indicated parameters. (Reproduced from Cleaver, 1967, with kind permission of the author.)...

Wittes and Kidwell (1973) have described a kinetic approach to measuring the pool size of dTTP which involves growing cells continuously in 10/iM tritiated thymidine which they found to label about 20% of the DNA thymine residues. They calculated that at this concentration 106 L929 cells growing in suspension would convert 9.5 pmol extracellular thymidine into dTTP per min during S-phase. 

When a single dose of tritiated thymidine is injected, all mitotic figures in the gastric pits are labeled within 48 hours (Fig. 3-12)." Labeling continues in succes-... 

---