Electron microscopy of structural detail in frozen biological specimens.

A procedure is described whereby preshadowed replicas can be obtained from frozen biological specimens which have been cut and then etched by sublimation of the ice from their surfaces. Electron micrographs showing details of the internal structure of plant virus crystals are presented to demonstrate the values of the procedure. Crystals of purified tobacco ringspot virus and squash mosaic virus and some portions of turnip yellow mosaic virus crystals have been shown to exhibit hexagonal packing. Sections through in situ crystals of tobacco mosaic virus show the rods to be parallel within each layer and arranged in a square net as viewed end on. Individual rods in each layer of the latter measure 300 mmicro in length and are somewhat tilted with respect to the rods of adjacent layers. This results in the formation of a herring-bone appearance when a crystal is cut perpendicular to its hexagonal face. It is suggested that the procedure outlined here might well serve to supplement other procedures for the preparation of many cytological specimens for electron microscopy.