Ultrastructural Studies on the Corneal Superficial Epithelium of Rats by in vivo Cryofixation with Freeze Substitution

Abstract

The method of in vivo cryofixation with freeze substitution (VC-FS) was used to examine the natural ultrastructure of the corneal superficial epithelium under the transmission electron microscope, and the results have been compared with the conventional method. With the VC-FS method, the ultrastructure down to 8 μm in depth was well preserved in the specimen. There was a layer of homogeneous material thought to be tear film on the corneal surface. Except for the intercellular spaces presenting under the first and sometimes also the second layers of the epithelial cells, other adjacent cells were in close contact without any spaces between them or between the desmosomes. In contrast, when using the conventional method, intercellular spaces were found between adjacent cells in all the layers of epithelium, and no tear film remained on the corneal surface. VC-FS is considered to be a better way to preserve the structure of tissue in its natural state than the conventional method and to be a useful method for the morphological study of the corneal epithelium and the tear film. Intercellular spaces, a kind of artifact, can be reduced by this method.