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Immunofluorescence analysis of human eye-banked corneas after applying low and high intensity collagen cross-linking

Poster Details

First Author: I.Beshtawi UK

Co Author(s):    C. O'Donnell   C. Hillarby   A. Brahma   F. Carely   H. Radhakrishnan  

Abstract Details



Purpose:

To compare the effects of low and high intensity collagen cross-linking (CXL) on the corneal collagen structure and stromal keratocyte distribution in normal human eye-banked corneas using immunofluorescence analysis.

Setting:

: Faculty of Life sciences, The University of Manchester, Manchester, UK.

Methods:

Ten postmortem human corneal pairs were studied. Five corneas were treated with the photosensitizer 0.1% riboflavin and UV-A irradiation (365nm, 3mW/cm2) for 30 minutes and the contralateral corneas were treated with riboflavin only and served as controls. In addition, five corneas were treated with 0.1% riboflavin and UV-A irradiation (365nm, 9mW/cm2) for 10 minutes, and the contralateral controls were treated with riboflavin only. Ultrathin histological sections from frozen samples were prepared and evaluated using immunofluorescence analysis (collagen type1 and DAPI) and imaged by fluorescence microscopy.

Results:

Better organisation of collagen fibers was noticed in the anterior stroma (down to 200┬Ám) of the low and high-intensity cross-linked cornea when compared with the controls. Moreover, fluorescence microscopy showed fewer keratocyte nuclei in the anterior stroma of the cross-linked corneas but with a more regular distribution than the controls in both cross-linked groups.

Conclusions:

Immunofluorescence microscopy demonstrated the effects of collagen cross-linking on eye-banked human corneas. This technique demonstrated keratocyte apoptosis and better collagen fiber organisation in the anterior part of the stroma of the cross-linked corneas. No difference between the low- and high-intensity cross-linked corneas was apparent which may imply similar efficacy of both protocols.

Financial Disclosure:

None

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