PRESERVATION AND STAINING OF MYELINATED NERVE-FIBERS

Six procedures are given for preservation of myelinated nerve fibers f or light or electron microscopic studies. These procedures fall into t wo main categories: those with and those without aldehyde fixation. Es sentially different effects are attained by application of tannic acid , saline, microwave or conventional heating, or a decreased temperatur e. All procedures end in osmication. Three main aspects of myelinated fiber morphology are taken into account when judging the quality of th eir preservation: axon, myelin sheath, and axon/myelin coherence. Each aspect can be preserved excellently, but always in combination with a less superior quality of the other two aspects. Superior myelin quali ty is attained using microwave irradiation, either with aldehydes to w hich tannic acid is added or without aldehyde fixatives. Superior axon quality is attained with aldehydes and (conventional) heating. Axon/m yelin coherence is best preserved by decreasing the temperature during the rinse with saline. Another two procedures provide good, though le ss superior, preservation of both axon and axon/myelin coherence. Next , the fixed tissue is embedded in plastic blocks from which semithin a nd ultrathin plastic sections are cut for light and electron microscop y, respectively. In addition to the standard procedure for toluidine b lue staining on semithin sections, two microwave-supported procedures are described, which can be used as alternatives if the staining resul t is unsatisfactory. Furthermore, a toluidine blue staining procedure is described for glycol methacrylate (GMA)embedded material, which can be used if larger sections are needed, (C) 1998 Academic Press.