Growth dynamics of mammalian cells monitored with automated cell cycle staining and flow cytometry

Abstract

To accurately observe high-frequency events during transient cell cycle kinetics, we have implemented a single step 15-min DNA staining protocol using automated flow cytometry. This protocol was used to sample and to analyze a Chinese hamster ovary cell culture for the DNA distribution, viable cell concentration, apoptotic cell concentration, and light scattering properties every 25 min over 4.5 days in response to a nutrient deprivation and a nutrient upshift. After the nutrient deprivation and exposure to fresh growth medium, two populations of cells started proliferating at different times likely corresponding to cells leaving the G0 and G1 cell cycle phases. After a nutrient upshift in late exponential growth, a cell cycle arrest occurred at the G1/S and G2/M boundary. The resulting cell cycle and proliferation kinetics followed damped oscillations that directly reveal the average time cells spend in each cell cycle phase. The observed detailed dynamics of the cell cycle progression is made possible through the high-frequency sampling enabled by automated flow cytometry. The approach should be useful in studying cell cycle perturbations in response to different environmental conditions resulting from exposure to specific nutrients or to drugs.