Expression of the prion protein gene (Prnp) and production of the PrP protein are essential requirements for acquisition and spread of transmissible spongiform encephalopathies such as Creutzfeldt-Jakob disease (CJD) in humans. Here we have developed an in situ hybridization method for use on human post-mortem central nervous system (CNS) tissues in order to determine those cell which are transcribing the Prnp gene and thus expressing PrP mRNA. Tissues from 11 adult individuals (age range 21-79 years) were analysed. Similar to previous studies in other animal systems, it was shown that PrP production occurs primarily in neuronal populations throughout the human brain. Neurones of the hippocampus, cortex, thalamus, cerebellum and medulla all synthesize PrP mRNA at readily detectable levels. No age-related differences were observed between the cases studied. It was also found that the ependymal cells produced PrP mRNA; these were the only non-neuronal cell type expressing the Prnp gene in the CNS. It is hoped that the information produced here will be helpful in understanding the pathology associated with CJD and other prion diseases in humans.