Brief Communication: Analysis of Cell Age Distribution During Ascites Tumor Growth 1,2

SUMMARY-Ascites lymphoma cells were inoculated into experimental mice. DNA distribution was serially de-termined by the flow microfluorimetry method during tumor growth.-J Natl Cancer Inst 53: 285-287, 1974. ASCITES LYMPHOMAS in experimental mice are a convenient system for the study of cell growth kinetics in the intact animal. After intraperitoneal (ip) inoculation of ascites lymphoma cells, a typical growth curve occurs, with an initial lag phase followed by a logarithmic increase in cell numbers and, finally, a stationary phase. Samples of tumor may be obtained at each stage of growth as a monodispersed, single cell suspension. We found that the LSA ascites tumor system of C57BL mice can be accurately ana-lyzed by tritiated thymidine uptake and DNA and RNA/cell (1). These trace the activities of the cell for synthesis of macromolecules concerned with cell replication, principally DNA. We now report deter-minations of DNA distribution made by the flow microfluorimetry (FMF) method. This method per-mits a direct analysis of cell age distribution in G I, S, and G2-M phases by DNA/cell content. FMF permits rapid and quantitative measurement of fluorescent light emission from fluorescently stained cells. It can measure directly the DNA distribution of many cells individually and, by means of output, dis-play data as a histogram of the various DNA/cell contents. A mammalian cell is normally diploid but will increase this DNA/cell content during DNA S phase to a tetraploid content. During G2 and early M, the cell will have a tetraploid DNA content and this will be reduced by mitosis to 2 diploid daughter cells. FMF measures DNA distribution and, by quantita-tion of this distribution, gives a direct measure of the proportion of cells in various phases of the cycle. For our purposes, a cell with a near-diploid content of DNA is designated as in GI phase; a cell with a doubling of the near diploid content is in G2-early M phase; and a cell with intermediate content, between GI and G2-early M, is in DNA S phase. The cell cycle terms, G I, G2, M, and S, were defined in (2). GO is a hypothetical, non proliferating state of cells. In these experiments, we tried to elucidate the nonproliferating stage of the advanced tumor, for a possible insight into the GO state