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RA Sack, S Sathe, A Beaton, B Bogart, OU Khan, P Asbell; Characterization of Proteins and Sialoglycoproteins in Normal and Dry Eye Tear Fluid . Invest. Ophthalmol. Vis. Sci. 2002;43(13):62.
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Purpose: To compare the protein and sialoglycoprotein (SG) profiles of tear fluid obtained from normals and individuals with dry eye syndromes. Methods: Tears were recovered from 29 individuals with no history of ocular surface tissue disease, 15 individuals with mild, 21 individuals with moderate and 15 individuals exhibiting severe aqueous deficiency dry eye syndrome. 5 minute after instillation of topical anesthetic, tears were recovered by Schirmer strip. A 5 mm wetted portion of the Schirmer strip which had not been in direct contact with the ocular surface tissue was extracted and separated in paralllel wiht a reflex tear satandard on 4-20 % SDS PAGE mini-gels under reducing conditions . Gels were reversibly Zn-imidazole stained, documented, destained and western blotted. Blots were cut horizontally at ∼100 kDa with the upper portion of the membrane probed using a mAb specific for a sialyl Lewis A epitope shown to exhibit a specificity similar to that of H185(Ab courtesy of Ilene Gipson)). The bottom portion of the membrane was reacted with Rb polyclonal Abs raised against human TSL. Blots were reacted with an alkaline phosphatase-linked secondary antibody with detection carried out using chemiluminescence. Following visualization the membranes were stripped and the bottom portions re-probed for albumin. Results:The SDS PAGE profile of a Schirmer strip collected tear samples differed from the profile of capillary tube collected reflex tear fluid in increased levels of albumin and an unidentified glycoprotein at ∼130 kDa decidely elevated in the Schirmer strip samples.The protein profiles of tear samples was similar in normals and those individuals who were diagnosed as having mild symptoms of either aqueous or lipid deficiency dry eye syndrome. In contrast, the profile from those subjects with severe to moderate dry eye syndrome was distinctly different. In most instances the concentration of inducible lacrimal specific tear proteins was markedly lower while the concentration of albumin was often elevated. Western blot analysis reveals that TSL was virtually absent or greatly reduced with epithelial cell derived SG reduced in a more variable manner. Conversely the concentration of albumin was often markedly increased. This relationship proved proportional to the severity of the pathology. Conclusion: Severe to moderate aqueous or combined aqueous/lipid deficiency dry eye syndrome is associated with an abnormal tear protein and SG profile.
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