Specimen preparation and chamber for confocal microscopy of the ex vivo eye

A chamber is described for maintaining the mechanical and physiological stability of the ex vivo eye during observation with confocal microscopy. The mechanical stability is provided by a plastic ring situated on the limbal region of the eye. The ring and supporting chamber are designed to reduce mechanical motion of the specimen. The ring and chamber size vary with the species and size of the eye under examination. The physiological stability over a period of approximately one hour is provided by immersing the eye in a bicarbonate Ringer's solution that is exchanged every five minutes. This fluid exchange is made between periods of microscopic observation. The suggested method for confocal microscopic observation of cornea and ocular lens in an ex vivo eye is to use a non-contact water immersion microscopic objective with a high numerical aperture. This is a non-invasive, non-applanating system for the confocal microscopical observation of ex vivo rabbit or human eye. Sample preparation and the specimen chamber are described. Optical sections of the cornea and lens obtained with a confocal microscope from a freshly removed ex vivo rabbit eye are presented as examples of applications of this technique.