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B.M. Ham, J.T. Jacob, M.M. Keese, R.B. Cole; Identification and Comparison of Proteins in Normal versus Dry Eye Rabbit Tears by MALDI–TOF MS . Invest. Ophthalmol. Vis. Sci. 2005;46(13):4426.
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Purpose: We investigated the use of matrix–assisted laser desorption/ionization time–of–flight mass spectrometry (MALDI–TOF MS) to compare the proteins in normal and dry eye rabbit tears. Methods: MALDI–TOF MS studies were performed on tear samples from normal and dry eyes of female New Zealand White rabbits. MALDI–TOF MS linear mode analysis of intact proteins in non–extracted tear fluid was performed using wax–coated Voyager plates and spot washing to determine the relative quantities of the tear proteins. Post source decay (PSD) and tandem mass spectrometry of the trypsin–digested peptides performed on two–phase extracted tear samples was used to identify the tear proteins. Experimental dry eye was induced by complete removal of the main and accessory lacrimal glands; tear break–up time (TBUT) values for the experimental dry eyes were 1/3 of those in the non–operated contralateral control eyes. Results: The lower molecular weight lipid binding proteins (approximately 10 kDa to 36 kDa) made up the largest proportion of the observed proteins, followed by the transferrins, which have higher molecular weights (70 kDa to 85 kDa). In general, the peaks associated with lipophilin CL 2 and lipocalin were increased in the dry eye tears, compared with normal tears from the non–operated contralateral control eyes. Conclusions: MALDI–TOF MS linear mode analysis of intact tear proteins is useful in identifying differences in expression of the lipid binding proteins in the low molecular weight range. Substantial differences in the amounts of lipid binding proteins in MALDI–TOF MS tear protein spectra from normal and dry eyes could indicate a disease state; such analysis may have an application in the diagnosis of dry eye.
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