Simultaneous visualization of neuronal protein and receptor mRNA

We describe a combined immunocytochemistry/in situ hybridization technique which allows for the simultaneous localization of protein and mRNA in a single cell. We have carried out these studies either on non-innervated skeletal myotubes or on myotubes which we have innervated with spinal cord explants or ciliary neurons. Our methods allowed us to detect acetylcholine receptor gene mRNA sequences which are expressed in low abundance within the cells and to determine the intracellular and intranuclear domains where these sequences are concentrated, as well as to identify neurons and their processes. The isotopic detection of RNA in combination with fluorescence microscopy produces high-resolution double-label images, with little background and good preservation of morphology, providing a powerful tool for detection of gene expression and protein content at the single-cell level.