DAPI fluorescence in nuclei isolated from tumors

In DNA histograms of some human solid tumors stained with nuclear isolation medium-4,6-diamidino-2-phenylindole dihydrochloride (NIM-DAPI), the coefficient of variation (CV) of the G(0)/G(1) peak was broad, and in nuclear volume vs DNA scattergrams, a prominent slope was seen. To determine the cause for this, nuclei from frozen breast tumors were stained with NIM-DAPI and analyzed after dilution or resuspension in PBS. In two-color (blue vs red) analysis, most of the slope and broad CV was due to red fluorescence of nuclei stained with NIM-DAPI, which was reduced on dilution or resuspension in PBS, resulting in elimination of the slope and tightening of the CV.