Proteomic assessment of tear fluid has become increasingly valuable in both human and veterinary research. The approach used to obtain tear samples may alter the resulting protein composition. This work investigates the tear protein patterns in dogs when samples are gathered using microcapillary tubes (MT), Schirmer tear strips (ST), and ophthalmic sponges (OS). Dog tears were obtained via MT, ST, and OS. Protein profiles were examined through two-dimensional electrophoresis (2-DE), and variations in the expression of individual protein spots were evaluated. Fourteen spots were subsequently identified by liquid chromatography–tandem mass spectrometry (LC-MS/MS). Total tear protein levels ranged from 2.80 to 4.03 μg/μL, with no significant differences detected among the three sampling techniques. Each collection method produced distinct protein-spot patterns in both number and intensity. MT, ST, and OS yielded 249, 327, and 330 spots, respectively. Albumin, haptoglobin, and lactoferrin derived from OS samples showed notably stronger spot intensities compared to the other methods. MT sampling resulted in reduced expression of nine proteins. These findings indicate that the technique used for harvesting tears influences proteomic outcomes. While ST remains widely used, it may not provide adequate detail for investigating specific tear proteins.
