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Global cell-by-cell evaluation of endothelial viability after two methods of graft preparation in Descemet membrane endothelial keratoplasty

Bhogal, M; Balda, MS; Matter, K; Allan, BD; (2016) Global cell-by-cell evaluation of endothelial viability after two methods of graft preparation in Descemet membrane endothelial keratoplasty. British Journal of Ophthalmology , 100 (4) pp. 572-578. 10.1136/bjophthalmol-2015-307534. Green open access

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Abstract

PURPOSE: To describe a novel method of global cell viability assessment for Descemet membrane endothelial keratoplasty (DMEK) and the comparison of two contemporary methods of donor tissue preparation. METHODS: DMEK transplants were prepared using two different methods: liquid bubble separation and manual peeling (n=8 each group). Samples were incubated with Hoechst, calcein-AM and ethidium homodimer prior to mounting on a curved imaging chamber. Z-stacked fluorescence microscopy images were combined to produce an in-focus global image capable of resolving all cell nuclei. Image processing software was used to define a calcein-positive live cell area, count all cell nuclei within this area and subtract ethidium-positive dead cells to derive the total viable endothelial cell count. Corrected global cell density was calculated by dividing the number of viable cells by the graft area, which had been corrected for imaging a curved surface. RESULTS: Corrected global cell density was lower than the central endothelial cell density in both groups: 85.5% of the pre-preparation central endothelial cell density in the peel group and 75.8% in the bubble group. Corrected global cell density was significantly lower in the liquid bubble separation group than in the peel group (p=0.04). CONCLUSIONS: Eye bank estimations of central endothelial cell density overestimate true cell density after graft preparation in DMEK. A peel method is less damaging and more consistent than a liquid bubble method. Cell loss correlated strongly with the degree of stromal hydration prior to bubble separation in the liquid bubble group.

Type: Article
Title: Global cell-by-cell evaluation of endothelial viability after two methods of graft preparation in Descemet membrane endothelial keratoplasty
Open access status: An open access version is available from UCL Discovery
DOI: 10.1136/bjophthalmol-2015-307534
Publisher version: http://dx.doi.org/10.1136/bjophthalmol-2015-307534
Language: English
Additional information: This is an Open Access article distributed in accordance with the terms of the Creative Commons Attribution (CC BY 4.0) license, which permits others to distribute, remix, adapt and build upon this work, for commercial use, provided the original work is properly cited. See: http://creativecommons.org/licenses/by/4.0/
Keywords: Cornea, Eye (Tissue) Banking, Treatment Surgery
UCL classification: UCL
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Brain Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Brain Sciences > Institute of Ophthalmology
URI: https://discovery.ucl.ac.uk/id/eprint/1476815
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