Tissue preparation for reconstruction of large-scale three-dimensional structures using a scanning electron microscope

A new tissue preparation method which gives scanning electron microscope images suitable for three-dimensional reconstruction of nerve tissue is described; the sample is embedded in an epoxy block which is ground, polished, and etched between taking successive cross-sectional images. Etching is done with a plasma produced by electron cyclotron resonance - a technique new to microscopy which gives minimal thermal damage and ensures clear images in which individual nerve fibres are easily resolved. Cropping of the sample is unnecessary, so that matching of laterally adjacent images is facilitated, and the method can deal with samples as large as 25 mm diameter x 20 mn depth.