Ultracryotomy and the corneal stroma

Williams, Leslie Rees (1984). Ultracryotomy and the corneal stroma. PhD thesis The Open University.

DOI: https://doi.org/10.21954/ou.ro.0000f91c

Abstract

Corneal stroma has two fundamentally interesting properties. Firstly, at physiological values of hydration, it is transparent to visible radiation. Secondly when bathed in aqueous solutions it will take up water and can achieve hydrations many times those of the physiological level with a consequent loss of transparency. These two important properties are intimately linked with two other features of corneal stroma. These are the lamellae of collagen fibrils of uniform diameter which make up the greater part of the stroma and the presence within the stroma of glycosaminoglycans which at physiological pH are negatively charged. The organisation of the collagen fibrils and their relationship to the glycosaminoglycans are important factors in understanding both the transparency of the tissue and the way in which it swells.

The work in this thesis falls into two parts. The first is concerned with the development of a technique for cutting ultrathin frozen sections of unfixed biological tissue for transmission electron microscopy. This technique, termed ultracryotomy, offers an alternative preparatory route for ultrastructural study in the electron microscope and in combination with X-ray microanalysis holds great promise for the identification and quantification of chemical distributions at an ultrastructural level. The second part describes experiments to measure the concentration of negatively charged poly-electrolytes within the stroma. This was done indirectly using a purpose built swelling pressure apparatus and then calculating a value for the charge concentration from a theory linking swelling pressure and charge concentration. As a comparison a direct measure of the charge concentration within the stroma was made using radioisotopes.

Exhaustive development was carried out with the technique of ultracryotomy and two fundamental limitations to the technique were established. These being the ultrastructural damage caused by the rapid freezing of the tissue and the artefacts produced when sectioning the frozen material. The results from the swelling pressure apparatus indicated that the value of charge concentration within the stroma was a function of the ionic strength of the stromal bathing medium, r The measures of charge concentration obtain- radioisotopically were considerably larger than those from the swelling pressure apparatus and showed no significant differences between the various bathing media. However there was a significant difference in the charge concentration between rapidly frozen and unfrozen corneal stromas as measured by this radioisotopic technique which suggests that rapid freezing was damaging the chemical integrity of the tissue.

Viewing alternatives

Download history

Metrics

Public Attention

Altmetrics from Altmetric

Number of Citations

Citations from Dimensions

Item Actions

Export

About

Recommendations