A method for comparing effects of different synchronizing protocols on mammalian cell cycle traverse. The traverse perturbation index.

Abstract

After treatment of Chinese hamster cells (line CHO) with various protocols for synchrony induction, the subsequent ability of cells to traverse the cell cycle (i e., to perform, an essential cell cycle process) has been determined by measurement of the DNA distribution pattern among cells in large populations with the Los Alamos flow microfluorometer In the cultures prepared by the various synchronizing techniques the vast majority of cells traversed the cell cycle in a normal fashion; however, in all cultures examined there remained small subpopulations which, though remaining viable for several days, could not carry out normal traverse. After reversible inhibition of DNA synthesis by means of a double-thymidine blockade, approximately 17% of the cells were unable to complete genome replication. After reversal of G(1) arrest resulting from cultivation of cells in isoleucine-deficient medium, 12 4% of the cells commenced synthesis of DNA but were unable to complete the S phase. Cells prepared by mitotic selection yielded a subpopulation (5 5% of the total cells) with a G(1) DNA content which remained viable but noncycling for at least 5 days. We propose a term "traverse perturbation index" which is defined as the fraction of cells converted to a noncycle-traversing state as the result of experimental manipulation. A knowledge of the perturbation index will allow direct comparison of effects on cell cycle traverse of various synchrony-induction protocols