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M. Ueffing, S. Hauck, M. Swiatek, C. Alge, S. Suppmann, S. Schöffmann, D. Pompetzki, C. Deeg, U. Welge-Lüssen, A. Kampik, T. Meitinger; Proteome Analysis of the Retina . Invest. Ophthalmol. Vis. Sci. 2003;44(13):2255.
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Purpose: Proteomic techniques allow for the visualization of complex protein expression patterns and identification of individual proteins by mass spectrometry. Methods: Four approaches to analyze aspects of retinal physiology will be presented, each based on high-resolution protein separation by two-dimensional electrophoresis, and protein identification by MALDI-TOF mass spectrometry. Results: 1: Proteome mapping: By comparative mapping of human, murine, bovine and avian (chicken) neural retinas by 2-D gel, we identified the 100 most abundant protein spots from a database generated in-house. Using a combination of biochemical and proteomic methods we have further analysed membrane proteins as well as protein-protein interaction patterns in mammalian photoreceptors. 2: Trans-differentiation in culture: Analysis of protein expression patterns from isolated primary retinal Müller glia cells in culture reveals the stability of protein expression during initial culturing, but shows loss of Müller glia cell-specific proteins, such as glutamate synthetase over time. The changes in expression reflect the trans-differentiation from a multifunctional, highly differentiated phenotype to a de-differentiated monolayer of cells in culture. 3: Secreted factors: Müller glia cells secrete factors that show a pronounced effect on photoreceptor survival by yet unknown means. A very sensitive method for searching secreted proteins from conditioned media using an in vivo 35-trans-S-label led to the detection of proteins secreted by Müller glia cells and by photoreceptors analyzed likewise. Proteins identified include alpha-1-antitrypsin, steroid monooxy-genase, collagens, clusterin, matrix metalloproteinase-2, tropomyosins and thrombospondin 1. 4: Identification of autoantigens for uveitis: Uveitis in horses is an autoimmune disease affecting the retina, as well as other parts of the eye. The identification of autoantigens triggering this disease is crucial for rational treatment. A technique using 2-D Gel separation, blotting, subsequent colloid gold staining and exposure of the blot to the affected horse’s serum led to identification of four autoantigens. Conclusion: Fields of applications for proteomic analyses of the retina but also some of their present limitations will be discussed.
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