Flow cytometry of isolated nuclei prepared from 9L rat brain tumor

Nuclei isolated by grinding 9L rat brain tumors gave a DNA distribution consisting of a 2C DNA peak due to noncycling normal cells and a bimodal distribution beginning at about 4C DNA due to the cycling tumor cells. Autoradiographic studies of nuclei from rats that received a pulse of 3H-TdR in vivo confirmed that the majority of proliferating cells were in the bimodal part of the distribution. The fraction of cells in S-phase determined from the DNA distributions was 15.3 per cent. This is consistent with a labeling index of 18 per cent determined from autoradiographs of tumor tissue and an S-phase fraction of 17 per cent determined from a fraction of labeled mitosis analysis (assuming that all of the 53 per cent noncycling cells are in G0). We also determined, by autoradiographic analysis of cells sorted from the tumor's mid-S-phase region, that only 58 per cent of these cells were labeled. We attribute this mostly to the presence of fluorescent debris from necrotic nuclei in this part of the distribution.