Insulin receptor mRNA was demonstrated in rat brain slices by in situ hybridization with three 35S-oligonucleotide probes and contact film autoradiography. Specificity was confirmed by showing that (a) excess unlabeled probe abolished the signal, (b) an oligonucleotide probe for rat neuropeptide Y mRNA showed a different distribution of hybridization signal, and (c) the distribution of insulin receptor binding was consistent with the distribution of insulin receptor mRNA. Insulin receptor mRNA was most abundant in the granule cell layers of the olfactory bulb, cerebellum and dentate gyrus, in the pyramidal cell body layers of the pyriform cortex and hippocampus, in the choroid plexus and in the arcuate nucleus of the hypothalamus.

Immunohistochemistry is widely used in biomedical research to localize specific epitopes of molecules in cells and tissues. The validity of interpretations based on immunohistochemistry requires appropriate positive and negative controls that are often not reported in publications. This omission may lead to incorrect interpretations and irreproducible results in the literature and contribute to wasted time, effort, and resources as well as erosion of confidence in scientific investigation by the general public, legislative bodies and funding agencies. The present article summarizes essential controls required for validation of immunohistochemical findings and represents a standard of practice for the use of immunohistochemistry in research and diagnostic investigations. Adherence to the guidelines described in the present article can be cited by authors as support for the validity of interpretations of the immunohistochemistry reported in their publications.