Characterization of human tear proteome using multiple proteomic analysis techniques

Tear proteome profiling may generate useful information for the understanding of the interaction between an eye and its contacting objects, such as a contact lens or a lens implant. This is important for designing improved eye-care devices and maintaining the health of an eye. Proteome profiles of tear fluids may also be used for disease diagnosis and prognosis. However, only a small volume of tear fluid (< 5 mu L) can be collected in a clinical laboratory under normal operational conditions, which makes proteome profiling a challenge. In this work we apply several proteomic analysis techniques, including gel-based and solution-based approaches with LC-ESI and LC-MALDI MS and MS/MS to gauge the relative merits of producing proteome profiles and to generate as broad a coverage of the tear proteome as possible from this small amount of sample. It is shown that a total of 54 proteins can be confidently identified using less than 5 mu L of tear fluid. Of these, 44 proteins can be detected by LC-MALDI MS alone with a consumption of 2 mu L of tear fluid. Furthermore, LC-MALDI can be used to determine post-translational modifications (PTMs), such as glycosylation and phosphorylation, without any sample enrichment or treatment. This work represents one of the most extensive proteome profiles (i.e., proteins identified and PTMs characterized) generated from tear fluids using clinically relevant amounts of sample.