Abstract
A scheme has been developed for deciphering the cell cycle time parameters of cell subpopulations that differ in their DNA ploidy level and that coexist in mixed heteroploid tumors. The S-phase analysis is presented. The approach is coupled to an automated imaging methodology for simultaneous determination of the Feulgen-stained DNA content and grain count of 3H-thymidine-labeled cells in autoradiographs (Sklarew RJ: J Histochem Cytochem 30:35, 30:49, 1982). The experimental designs involve 3H- and 14C-thymidine double labeling and Colcemid incubation. The deconvolution of the S-phase ploidy composition is illustrated in rat sarcoma cultures comprising four major ploidy subpopulations; with G-2 and mitotic DNA contents of approximately 4C, 8C, 16C, and 32C. The components were identified by their DNA ploidy level, and their S frequencies and labeling indices were obtained. A scheme is also developed and validated for obtaining the S-phase emptying profile of component ploidy subpopulations, and their cell flux at the S/G-2 and G-2/mitosis phase boundaries. In the sarcoma cultures S mobility was found to decrease with increasing DNA ploidy level over the entire ploidy range.