We have used immunohistochemical techniques and a monoclonal antibody against proliferating cell nuclear antigen (PCNA) to investigate the proliferative activity of glial cells in mice with experimental Creutzfeldt-Jakob disease (CJD), and in human cases of CJD, kuru and Gerstmann-Sträussler-Scheinker syndrome (GSS). Only a small proportion of hypertrophic astrocytes showed PCNA immunoreactivity (labelling index, LI: 0-4.5%). PCNA-specific immunostaining was confined entirely to cell nuclei. During the early stages of illness, with minimal CJD pathology, PCNA-immunopositive nuclei were occasionally observed in the subependymal zone of experimentally infected mice. From 18 weeks postinoculation, PCNA-immunopositive astrocytes were most frequently found in the corpus callosum and cerebellar white matter; regions which characteristically exhibit robust vacuolation. No other cells, particularly no cells of microglial morphology, showed PCNA immunoreactivity. In human cases of kuru, CJD and GSS, no PCNA-immunopositive cells were detected despite the presence of numerous microglial cells and reactive hypertrophic astrocytes. These results indicate that only a limited proportion of astrocytes proliferate in the experimental models of subacute spongiform encephalopathies and that microglia are probably postmitotic cells.