Ultrastructural Studies of Collagen Fibers of the Cornea and Sclera by a Quick-Freezing and Deep-Etching Method

The ultrastructure of collagen fibers of the cornea and sclera of rabbits was studied by a newly developed quick-freezing and deep-etching method. An isopentane-propane mixture (––193°C) cooled in liquid nitrogen was used for quick-freezing of the tissues. The frozen tissues were fractured in nitrogen, deeply etched and shadowed with platinum and carbon. Corneal collagen fibers were observed to be longitudinally arranged and separated by moderately wide interfibrillar spaces. Interconnecting filaments were three-dimensionally radiating between the collagen fibers, which were 40.6 ± 5.0 nm in diameter and 42.0 ± 2.8 nm in periodicity. In contrast, scleral collagen fibers were compactly organized, with few interconnecting filaments. Five kinds of striations were clearly observed on the collagen fibers, which were 217.3 ± 50 nm in diameter and 67.5 ± 1.4 nm in major periodicity. These striations could hardly be observed in the fractured scleral collagen fibers. The present study reveals the ultrastructural differences between corneal and scleral collagen fibers through the use of a quick-freezing and deep-etching method.