A Staining Method for Sections of Osmium-Fixed, Methacrylate-Embedded Tissue

The increasingly broad application of electron microscopy to the study of cytological detail in both normal and pathological tissues has created a need for an improved staining method for light microscopy of material processed primarily for electron optical investigation. A staining technic that would produce results comparable to those obtained with hematoxylin and eosin should prove to be of value in correlating existing histological information with the findings of electron microscopy. Also, many problems of interpretation can best be approached by a comparison in the light and electron microscopes of adjacent sections through individual structures (3). Such a comparison is made difficult by the fact that the fixative of choice for electron microscopy (osmium tetroxide) alters or destroys the affinities of tissues for ordinary stains. Furthermore, it seems likely that additional loss of stainability may occur as a result of oxidative processes that accompany methacrylate embedding (1). Houck and Dempsey (2) and others have utilized certain elastin and collagen staining methods for light microscopy of osmium-fixed, methacrylateembedded tissues. In our laboratory the application of such stains as the periodic acid-Schiff reaction, ammoniacal silver, aldehyde fuchsin, various hematoxylin solutions, azocarmine, and a variety of "counterstains" have not consistently yielded highly satisfactory results. None of these stains alone or in combination approach the quality of microscopic differentiation achieved with classical histologic fixation and embedding techniques. Adequate demonstration of stained tissue by black and white photography is difficult and is achieved only with special photographic methods. The purpose of this communication is to outline a staining method, currently in use for special problems in light microscopy, which we have