May 2004
Volume 45, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2004
Proteome analysis of normal human corneas
Author Affiliations & Notes
  • T. Moller–Pedersen
    Department of Ophthalmology, Aarhus University Hospital, Aarhus, Denmark
  • I.B. Thogersen
    Department of Molecular Biology, University of Aarhus, Aarhus, Denmark
  • H. Karring
    Department of Molecular Biology, University of Aarhus, Aarhus, Denmark
  • G.K. Klintworth
    Department of Pathology, Duke University Medical Center, Durham, NC
  • J.J. Enghild
    Department of Molecular Biology, University of Aarhus, Aarhus, Denmark
  • Footnotes
    Commercial Relationships  T. Moller–Pedersen, None; I.B. Thogersen, None; H. Karring, None; G.K. Klintworth, None; J.J. Enghild, None.
  • Footnotes
    Support  NIH Grant RO1–EY12712
Investigative Ophthalmology & Visual Science May 2004, Vol.45, 3517. doi:
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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)

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Abstract

Abstract: : 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.

Keywords: cornea: stroma and keratocytes • proteomics • cornea: basic science 
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