The effect of fixation on corneal endothelial cell dimensions and morphology in scanning electron microscopy

Soft tissue specimens shrink during fixation, dehydration and critical point drying when prepared for scanning electron microscopy (SEM). This can cause serious artifacts not only in 'compact' tissues but especially in hollow structure, like the eye, where the chambers are lined by delicate layers such as the corneal endothelium. In this study various glutaraldehyde and formaldehyde fixations at different concentrations with or without 5% sucrose were tested. Dimensional as well as morphological changes of rabbit corneal endothelial cells were evaluated. The mean surface diameter and area of rabbits' central corneal endothelial cells were measured first in vivo by specular microscope. Thereafter the same corneas were fixed in 9 different solutions and processed for scanning electron microscopy. The surface structure of the same endothelium was then photographed with SEM. The cell dimensions were remeasured. According to our results 1.25% or 2.50% glutaraldehydes without sucrose gave the best surface preservation and caused the least shrinkage.