Sample preparation techniques for conventional and high resolution scanning electron microscopy of the central...

In the present paper some basic sample preparation techniques for scanning electron microscopy of nervous tissue are described. These basic preparative methods include conventional scanning electron microscopy or slicing technique, ethanol-cryofracturing technique, freeze-fracture method using either liquid nitrogen (slow freezing) or Freon 22 cooled by liquid nigrogen (fast freezing), improved freeze-fracture method with delicate specimen preparation and chromium coating, ultrasonic microdissection, and "creative tearing" technique. Some basic principles, advantages and limitations are critically considered. In addition, some specific applications in neurobiological research are reported. Emphasis is placed upon understanding the sources and nature of artifacts that are likely to be produced in each preparatory step. Examples are given of the results obtained with the different types of nerve tissue preparation, using the cerebellar cortex as a model of the central nervous system. According to the author's experience, the slicing technique is recommended for studying cytoarchitectonic arrangement of gray centers, the ethanol-cryofracturing technique for tracing short intracortical circuits, and the freeze-fracture methods for analysis of nerve cell cytoplasmic and nuclear compartments. An attempt is made to explain results obtained in relation with nerve cell biology.