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P.J. Anderton, S.T. Tragoulias, M. Ball, F. Miano, G.R. Dennis, T.J. Millar; Purified Bovine Ocular Mucin Has Virtually No Surface Activity . Invest. Ophthalmol. Vis. Sci. 2003;44(13):2502.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: It is commonly believed that ocular mucins are surface active in tears. We have further investigated this idea by purifying bovine ocular mucin and comparing its surface activity with commercially available bovine submaxillary mucin (BSM), purified BSM, and hyaluronic acid. Methods: Ocular mucin was extracted from fresh bovine conjunctiva using 6M urea, 0.1% SDS, and 10mM DTT at 4°C for 16h. The extract and commercial BSM were purified using a Pharmacia Superose 6 column (16x600mm) in 50mM Na phosphate buffer containing 1M NaCl and 3mM DTT at pH7.5. It was desalted into water using a Pharmacia Fast Desalt column (10x100mm) and freeze dried. Purified and unpurified mucins were checked for purity using SDS-PAGE. The different mucins and hyaluronic acid were applied to an air/phosphate buffered saline interface in a Langmuir trough (maximum surface area 80cm2). The surface pressure was measured using a Wilhelmy balance as the film was compressed and expanded. Results: Ocular mucin and purified BSM had no detectable bands below 200kD. Commercial BSM had multiple bands with strongly stained bands at ~60kD, 20kD and 10kD. Bovine ocular mucin (10mg/mL, 50µL) and hyaluronic acid (1mg/mL, 45µL) had no detectable surface activity. Purified BSM (1mg/mL, 20-1000µL) gave negligible surface activity, but at a higher concentration (5mg/mL, 200µL) showed a maximum surface activity of 25mN.m–1. Commercial BSM gave strong surface activity even with low concentrations and low volumes (1mg/mL, 20µL). Conclusions: By examining the surface activity of mucin and hyaluronic acid, there is strong evidence to suggest that neither ocular mucin nor hyaluronic acid have surface activity, and hence any contribution to the surface activity of tears must come from their interaction with other tear film components. These differ from commercial BSM which had marked surface activity. This indicates that unpurified BSM cannot be used as a substitute for ocular mucin when modelling the tear film.
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