Protease-elicited TUNEL positivity of non-apoptotic fixed cells

The appearance of free DNA ends in the chromatin is usually considered an i ndication of advanced apoptosis. Unexpectedly, the nuclei of non-apoptotic cells derived from mouse thymuses could be specifically labeled by terminal transferase after proteinase K treatment of the fixed, cytocentrifuged sam ples. Artifactual mechanical or contaminating nucleolytic factors have been ruled out as players in the generation of free DNA ends. The phenomenon wa s detected in both formaldehyde- and ethanol-fixed specimens, in agarose-em bedded fixed cells, and in chromatin spreads. By urea-agarose gel electroph oresis, the average single-strand size of the DNA molecules carrying the fr ee ends was found between 50 and 250 kb. We suggest that ss discontinuities preexisting in the fixed normal cells are unmasked by protease treatment e liciting TUNEL (terminal transferase-mediated nick end-labeling) positivity .