Unembedded, aldehyde-fixed tissue, sectioned for transmission electron microscopy

Abstract

It has been demonstrated that it is feasible within limits to section a wide variety of unembedded, glutaraldehyde-fixed tissues thinly enough for transmission electron microscopy. Much cytological detail is preserved. The success of the method is thought to depend mainly upon glutaraldehyde adequately cross-linking cytosol proteins so that cellular components are not only held in place, but the tissue is sectionable after it is air dried. A polyvinyl-acetate emulsion in the form of carpenter's white glue is used as an external support for tissue blocks. If necessary, this can be rendered insoluble by the addition of a small quantity of serum albumin, subsequently cross-linked with glutaraldehyde. Glass knives are advisable for sectioning, and sections are floated on a layer of water reaching the knife edge with a minimal meniscus. Since alkaline lead stains have proven to be quite destructive to unembedded tissue, and other positive stains have not been found that are particularly effective, partially neutralized phosphotungstic acid, used as a negative stain, is the principal staining technique employed. Because it is thought that the technique ultimately may prove to be most useful for immunocytochemical applications, emphasis has been placed upon studies of tissues fixed only with glutaraldehyde. If tissue is secondarily fixed with osmium tetroxide and/or treated with uranyl salts, however, tissue blocks then have superior sectioning properties, and can be more effectively and positively stained.