An improved silver staining technique as an alternative nuclear or combined nuclear nerve-fiber impregnation for comparative light-, secondary and backscattered electron scanning microscopy.

Slices of rat brain were stained by a new silver impregnation technique. This method takes into consideration the pH-dependent differences of silver stain affinity of nerve tissues and can be used alternatively as a stain for nuclei or as a method for combined demonstration of nuclei nerves fibers. The slices were studied at the light microscopical (LM) level and subsequently with a scanning electron microscope, using secondary (SSEM = classical SEM), and backscattered electron detectors (BSEM). This new silver staining technique offers the opportunity of comparative studies with regard to different information acquired with LM, SSEM and BSEM. The described method allows to distinguish between nervous and glial tissue without necessarily damaging the glial tissue surrounding the nerve fibers. Specifically, scanning electron microscopy with backscattered electron detector of in situ preparations provides a higher contrast of stained and unstained tissue and increased depth of focus as compared to secondary electron detectors.