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P. Meyer, X. Xin; Primary Cell Culture of Meningothelial Cells - A New Model for Future Studies of Optic Nerve Diseases. Invest. Ophthalmol. Vis. Sci. 2008;49(13):2304.
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In a previous investigation we showed that the occurrence and amount of meningothelial cell nests in the subarachnoid space were significantly increased in histological examinations of glaucoma. For further investigation of a possible role of meningothelial cells in the pathogenesis of optic nerve diseases in vitro, pure meningothelial cells are necessary. We have therefore developed a culture method of meningothelial cells from the arachnoid layer covering the optic nerve.
Meningothelial cells scraped from the arachnoid layer of optic nerve were cultured for 2-3 weeks. The primary passage of cells was purified using magnetic beads to remove fibroblasts from the cultured cells. After cells were detached by 0.05% Tripson-EDTA, anti-fibroblast microbeads were added in cell suspension and applied onto the magnetic column. Fibroblasts labeled with anti-fibroblast micro-beads were collected by magnetic column, and the effluent that passed through the column was the collection of the unlabelled cell fraction. The immunofluorenscence and flow cytometry were done to confirm that the unlabelled cells were meningothelial cells by using keratin sulfate antibody which was visualized with fluorescence Alexa-fluor conjugated goat-anti-mouse secondary antibody.
Most cells were positive staining for keran sulfate in immunoflurorescence and at least 90% cells were keratin sulfate-positive meningothelial cells in flow cytometry after the separation by using anti-fibroblast microbeads.
Our methods for primary meningothelial cell culture from arachnoid layer of optic nerve combining the method of magnetic beads for removing fibroblasts is effective in obtaining pure and rich meningothelial cells.
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