CELL MEMBRANE-DEPENDENT CHROMATIN CONDENSATION

It is shown by means of now cytometry that during several seconds afte r cell membrane damage by a non-ionic detergent in physiologically rel evant buffer solution, the chromatin of mouse thymocyte nuclei undergo es a drastic decondensation, which is revealed by a sharp increase of binding of DNA-specific fluorochromes (olivomycin or propidium iodide) and of DNA accessibility to DNAse I digestion. A similar change is ob served in dead cells. Roughly half of this decondensation can be preve nted by lowering the pH of the outside medium to the level known to be inside the cells; the other half remains thus far unexplained (divale nt cations and the difference between small anion species seem not to be involved). The approach is based on a novel observation that fixati on by formaldehyde conserves chromatin structure before the action of detergent. Flow cytometric assay is proposed for monitoring these cond ensation/decondensation events in media of different composition. In a ddition, a new approach to viable/dead cell determination, which has t he advantage of immediately fixing the cell state and preserving it fo r a reasonably long time, is proposed. (C) 1995 Wiley-Liss, Inc.