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Basilio Colligris, Alba Martin-Gil, Begoña Fonseca, Gonzalo Carracedo, Jesus Pintor, Bioquimica Farmacologica Del Ojo-OcuPharm; Improving 3D mucin visualization by a new device to make impression cytology comparing normal and muco-deficient subjects. Invest. Ophthalmol. Vis. Sci. 2014;55(13):4874.
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The aim of this study was to validate the use of a new medical device to make impression cytology with a new mucin visualization technique allowing not only assess Goblet cells density, but to quantify mucins release in the ocular conjunctiva.
In this study were taken conjunctival impression cytology samples from 25 healthy subjects (25 eyes) and samples from 29 subjects (29 eyes) with some kind of muco-deficient pathology. It was used the Eyeprim TM device that requires no topical anesthesia and allows collection of 50 % more cells than the traditional method. Treatment of cytology samples was done by PAS- hematoxylin. For measuring cell density, the display of cytology by impression was made by transmitted light microscopy. For the new method of displaying mucins in 3D, the samples were analyzed with a confocal laser microscope (LSM) by which the different sample dyes were excited at 488 and 543 nm. For each image were made a series of optical slides in axis z, that were reconstructed to obtain a three dimensional image.
With this new technique the Goblet cells are easily distinguishable from the epithelial cells, but also the 3D image permits to measure new parameters, such as the height of the mucin cloud secreted by each cell ( 8.81 ± 4.00 microns ), the mucin diffusion of the epithelial cells ( 2.77 ± 1.00 microns ) and the thickness of the Goblet cell. These parameters were significantly decreased, by about 70 % and 40% respectively, in the group of muco-deficient patients.
The mucin visualization technique by confocal laser microscopy in 3D is a solid diagnostic tool, providing a number of parameters related to both Goblet cells secretion and the cell itself, allowing more objective assessments of the mucin secretion status, than the classical measure of cell density. The use of the new medical device for impression cytology EyePrim improves the features of 3D LSM analysis.
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