Improved ultrastructural preservation of rat ciliary body after high pressure freezing and freeze substitution: A perspective view based upon comparison with tissue processed according to a conventional protocol or by osmium tetroxide/microwave fixation

Abstract

Conventional fixation of the delicate, highly folded rat ciliary body and its iridial extension, as well as of vitreal structures, is associated with the induction of a number of artifacts, thus limiting the reliability of morphological interpretations. Improved ultrastructural preservation may be achieved by microwave heating in combination with osmium tetroxide fixation. This protocol, although simple and cheap, yields results, particularly with respect to the extracellular matrix compartment between inner and outer ciliary epithelial cells, which are not greatly inferior to those obtained by implementing the sophisticated high pressure freezing and freeze substitution technique. The latter affords good to very good ultrastructural preservation of epithelium and stromal components, such as blood vessels, neural elements, smooth muscle cells, fibrocytes, and free cells, up to a depth of 50–100 μm from the tissue surface. Its superiority over osmium tetroxide/microwave fixation is revealed in the cytoplasmic, intraorganellar, and vitreal matrix compartments, which incur no obvious losses.