Abstract
Hoechst 33342 can be used for flow microfluorimetry (FMF) analysis of the DNA content of living Chinese hamster cells, giving good resolution (coefficients of variation (CVs) 6%) with relatively nontoxic staining regimens. The dye is, however, a very efficient inhibitor of DNA synthesis, with a marked depression of the DNA synthetic rate for V79 cells observed at concentrations tenfold less than those required for optimal FMF resolution. Nontoxic and minimally toxic Hoechst concentrations also resulted in demonstrable mutation, as assayed by 6-thioguanine resistance. Hoechst disappearance from cells returned to normal growth medium after staining suggests two components of binding, since about half of the total stain is rapidly removed, whereas the rest is apparently diluted only by cell division. Cells containing Hoechst 33342 die more rapidly than control cells when held at 4 degrees C and are also more susceptible to inactivation by the ultraviolet laser beam if operated at approximately greater than 100 mW power. Thus, Hoechst 33342 can provide information about the DNA content of living cells, but at the expense of minimal toxicity, moderate mutation, and significant cell cycle perturbations.