Enhancement of immunoperoxidase staining using osmium tetroxide

A modification of the peroxidase-antiperoxidase immunohistochemical method involving the use of OsO4 for the visualization of antigens in tissue using thick sections is described. This modification has lead to consistent enhancement of the intensity of the reaction product and thus to a further amplification of sensitivity in this particular system. The present procedure results in a sensitive demonstration of neuronal antigens such as peptides and synthesizing enzymes in neurons. The method allows the investigator to analyze the cytoarchitecture and geometry of these neurons and their projections in the nervous system as well as more easily detect single, weakly stained nerve fibers. In 50 micrometers sections, immunopositive processes can be followed over long distances. In the present study, the catecholamine-synthesizing enzyme, phenylethanolamine-N-methyltransferase (PNMT), and the neuropeptide, somatostatin, have been used as examples from the central nervous system of the rat, but the procedure is applicable to other antigens as well as other cell types and tissues.