Differential gel electrophoresis of the tear proteome

Markoulli, M; Papas, E; Cole, N; Holden, B

Differential gel electrophoresis of the tear proteome

Markoulli, M Papas, E Cole, N

Holden, B

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Publication Type:: Journal Article
Citation:: Optometry and Vision Science, 2012, 89 (6)
Issue Date:: 2012-06-01

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Field	Value	Language
dc.contributor.author	Markoulli, M	en_US
dc.contributor.author	Papas, E	en_US
dc.contributor.author	Cole, N https://orcid.org/0000-0002-9866-4694	en_US
dc.contributor.author	Holden, B	en_US
dc.date.issued	2012-06-01	en_US
dc.identifier.citation	Optometry and Vision Science, 2012, 89 (6)	en_US
dc.identifier.issn	1040-5488	en_US
dc.identifier.uri	http://hdl.handle.net/10453/37806
dc.description.abstract	Purpose. This study evaluates the use of differential gel electrophoresis (DIGE) for tear analysis and applies this technique to establish the effect of extended contact lens wear on the tear proteome. Methods. Flush tears were collected from nine healthy non-contact lens wearers at baseline, during the first day, and after 1 month of contact lens wear. Participants wore AIR OPTIX on an extended wear (EW) schedule. Tears were collected at mid-day and upon waking and analyzed for concentrations of total protein using the bicinchoninic acid assay method. DIGE was then performed to detect biomarkers likely to be affected by contact lens wear. Technique variability, physiological variability, and the effect of EW were established. Proteins with significantly changed abundance during lens wear were cut and identified with mass spectrometry. Results. Both Cy3 and Cy5 identified the same proteins relative to Cy2 with the highest fold difference of 4.4. The proteome varied by 3% at the same time point, with a significant increase in protein spots found between mid-day (257) and upon awakening (298). With EW, of the 311 proteins spots identified upon awakening, 15 differed significantly. Of the five differences between baseline and the first night of EW, all were greater at baseline. Of the differences between baseline and 1 month, six were greater at 1 month and four were greater at baseline. Conclusions. DIGE is a repeatable proteomic technique capable of determining differences in the tear proteome. The extent of the participants' experience with lens wear is an important factor to consider when analyzing the tears from lens wearers, as neophytes appear to differ from those with experience. © 2012 American Academy of Optometry.	en_US
dc.relation.ispartof	Optometry and Vision Science	en_US
dc.relation.isbasedon	10.1097/OPX.0b013e318255dc46	en_US
dc.subject.classification	Ophthalmology & Optometry	en_US
dc.subject.mesh	Tears	en_US
dc.subject.mesh	Humans	en_US
dc.subject.mesh	Proteome	en_US
dc.subject.mesh	Electrophoresis	en_US
dc.subject.mesh	Reproducibility of Results	en_US
dc.subject.mesh	Contact Lenses, Extended-Wear	en_US
dc.subject.mesh	Reference Values	en_US
dc.subject.mesh	Adult	en_US
dc.subject.mesh	Female	en_US
dc.subject.mesh	Male	en_US
dc.title	Differential gel electrophoresis of the tear proteome	en_US
dc.type	Journal Article
utslib.citation.volume	6	en_US
utslib.citation.volume	89	en_US
utslib.for	11 Medical and Health Sciences	en_US
pubs.embargo.period	Not known	en_US
pubs.organisational-group	/University of Technology Sydney
pubs.organisational-group	/University of Technology Sydney/Faculty of Science
pubs.organisational-group	/University of Technology Sydney/Faculty of Science/School of Mathematical and Physical Sciences
utslib.copyright.status	closed_access
pubs.issue	6	en_US
pubs.publication-status	Published	en_US
pubs.volume	89	en_US

Abstract:

Purpose. This study evaluates the use of differential gel electrophoresis (DIGE) for tear analysis and applies this technique to establish the effect of extended contact lens wear on the tear proteome. Methods. Flush tears were collected from nine healthy non-contact lens wearers at baseline, during the first day, and after 1 month of contact lens wear. Participants wore AIR OPTIX on an extended wear (EW) schedule. Tears were collected at mid-day and upon waking and analyzed for concentrations of total protein using the bicinchoninic acid assay method. DIGE was then performed to detect biomarkers likely to be affected by contact lens wear. Technique variability, physiological variability, and the effect of EW were established. Proteins with significantly changed abundance during lens wear were cut and identified with mass spectrometry. Results. Both Cy3 and Cy5 identified the same proteins relative to Cy2 with the highest fold difference of 4.4. The proteome varied by 3% at the same time point, with a significant increase in protein spots found between mid-day (257) and upon awakening (298). With EW, of the 311 proteins spots identified upon awakening, 15 differed significantly. Of the five differences between baseline and the first night of EW, all were greater at baseline. Of the differences between baseline and 1 month, six were greater at 1 month and four were greater at baseline. Conclusions. DIGE is a repeatable proteomic technique capable of determining differences in the tear proteome. The extent of the participants' experience with lens wear is an important factor to consider when analyzing the tears from lens wearers, as neophytes appear to differ from those with experience. © 2012 American Academy of Optometry.

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