To determine whether cultured astrocytes express opioid gene mRNA, a method was developed for co-localizing a cell-type specific immunocytochemical marker for astrocytes, glial fibrillary acidic protein (GFAP), and proenkephalin mRNA in situ hybridization signal using high affinity cRNA probes. GFAP immunoreactivity and proenkephalin mRNA hybridization reaction were examined in intact glial cell preparations from neonatal mice that were cultured for 4-6 days prior to fixation. The double labelling method described herein permits the unambiguous identification of mRNA expression in specific populations of intact cultured cells using cell type-specific markers.