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T. Moller–Pedersen, I.B. Thogersen, H. Karring, G.K. Klintworth, J.J. Enghild; Proteome analysis of normal human corneas . Invest. Ophthalmol. Vis. Sci. 2004;45(13):3517.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: To determine the protein expression profile of normal human corneas and thus establish a baseline for further studies of pathological specimens. Methods: Thirty normal human corneas (7–mm central trephination) containing all layers were dried and homogenized. To identify as many proteins as possible, three different extraction protocols were used: 1) extraction in urea/detergent buffer followed by 2D gel electrophoresis at various pH gradients and MALDI mass spectrometry of all gel spots; 2) extraction in SDS sample buffer followed by SDS–PAGE; the gel lane was sliced in 2 mm strips, digested with trypsin and analyzed by LC–MS/MS; and 3) treatment with CNBr/trypsin followed by separation of fragments by ion exchange and analysis using LC–MS/MS Multidimensional protein identification technology (MudPIT). Results: Numerous proteins were identified including collagens, proteoglycans, protease inhibitors, antioxidants, angiogenesis inhibitors, crystallins, lipoproteins, chaparons, and metabolic proteins. Several proteins not previously identified in the cornea were detected. The soluble fraction was dominated by albumin, TGFBIp (bigh3), and immunoglobulins. Conclusions: The identified proteins provide a reference library for targeted studies of corneal transparency and diseases.
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