Flow cytometric analysis of DNA content of living and fixed cells: a comparative study using various fixatives

The majority of studies dealing with DNA analyses are made on fixed cells. In this context, the efficiency as fixatives of ethanol, methanol, acetone, Carnoy, Boehm-Sprenger and aldehydes was determined using two different DN A fluorescent probes, Hoechst 33342 and propidium iodide. The purpose of ou r study was to find the fixative that would provide the best results with r espect to the following parameters: aggregates, cell size and granularity, and DNA staining analysis. Using murine fibroblasts, we found that 68% etha nol, 85% methanol and aldehydes did not increase aggregate formation, where as Carnoy, acetone or Boehm-Sprenger fixatives did. The results show that a ldehydes seem to alter cell size least. All fixatives induce an increase in cell granularity, which is very pronounced with alcohols, but aldehydes al ter morphology less than alcohols. We observed that the fixatives giving th e best resolution with Hoechst 33342 staining lead to a lower measurement v ariability than with propidium iodide staining. This study leads us to conc lude that 68% ethanol and 85% methanol can be considered as appropriate fix atives for flow cytometry studies of DNA content. (C) 1998 Chapman & Hall.