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N. Gandhi, K. Goins, R. Mullins, G. S. Enriquez; The Effect of Shipping on Endothelial Cell Viability After Laser Assisted Keratoplasty Donor Preparation. Invest. Ophthalmol. Vis. Sci. 2009;50(13):630.
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Laser assisted keratoplasty (LAK) using the femtosecond laser provides more wound surface area, increased wound integrity, and decreased suture retention time. The use of pre-cut LAK tissue prepared by eye banks may also reduce surgeon stress and improve operating room efficiency. The purpose of this study was to determine using two assays whether shipping pre-cut LAK tissue compromises endothelial cell viability.
Part One: Eight recently enucleated corneas prepared by the Iowa Lion’s Eye Bank (ILEB) with the femtosecond laser using a zig-zag wound configuration were shipped overnight. Four buttons were left attached to the corneo-scleral rim and four were shipped free-floating in Optisol GS. 24 hours after LAK, the corneal endothelium was treated with trypan blue dye and rinsed in phosphate buffered saline. Digital images were converted to bitmap files using Adobe Photoshop. The histogram function of ImageJ software was used to quantify white and black pixels corresponding to live or dead endothelium. Significance was determined using a paired Student’s t test.Part Two: Ten recently enucleated corneas were prepared by the ILEB with the femtosecond laser using top-hat (n =4), zig-zag (n = 4) and mushroom (n=2) configurations and were shipped overnight. Half of the specimen from each group were shipped free-floating in Optisol GS and half from each group were shipped attached to a corneo-scleral rim. 24 hours later, corneas were stained using the LiveDead Assay Kit (Invitrogen). The central 10x field of each specimen was photographed using a rhodamine cutoff filter on an Olympus BX41 fluorescence microscope. Red nuclei indicating dead cells were counted; significance was determined using a paired Student’s t test.
Part One: The mean endothelial cell death in the attached group was 0.044%, and in the free group was 0.12%. There was no statistically significant difference in endothelial cell viability between the free floating and attached groups (p = 0.395).Part Two: The mean endothelial cell death in the attached group was 0.024 cells/mm2 and in the unattached group was 0.025 cells/mm2 ; there was no statistically significant difference between these two groups (p= 0.877). The mean cell death in the top-hat group was 0.032 cells/mm2 and in the zig-zag group was 0.025 cells/mm2 ; there was no statistically significant difference between these two groups (p = 0.644).
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