Please use this identifier to cite or link to this item: http://hdl.handle.net/1959.14/42511
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- Title
- Comparing SILAC and two-dimensional gel electrophoresis image analysis for profiling urokinase plasminogen activator signaling in ovarian cancer cells
- Related
- Journal of proteome research, Vol. 6, Issue 6, p.2105-2112
- DOI
- 10.1021/pr060638v
- Publisher
- American Chemical Society
- Date
- 2007
- FoR/RFCD Code(s)
-
030199 Analytical Chemistry not elsewhere classified
030406 Proteins and Peptides
- Author/Creator
- Uitto, Pauliina M
- Author/Creator
- Lance, Braddon K
- Author/Creator
- Wood, Graham R
- Author/Creator
- Sherman, James
- Author/Creator
- Baker, Mark S
- Author/Creator
- Molloy, Mark P
- Description
- Two-dimensional gel electrophoresis (2-DE) image analysis is conventionally used for comparative proteomics. However, there are a number of technical difficulties associated with 2-DE protein separation that limit the depth of proteome coverage, and the image analysis steps are typically labor-intensive and low-throughput. Recently, mass spectrometry-based quantitation strategies have been described as alternative differential proteome analysis techniques. In this study, we investigated changes in protein expression using an ovarian cancer cell line, OVMZ6, 24 h post-stimulation with the relatively weak agonist, urokinase-type plasminogen activator (uPA). Quantitative protein profiles were obtained by MALDI−TOF/TOF from stable isotope-labeled cells in culture (SILAC), and these results were compared to the quantitative ratios obtained using 2-DE gel image analysis. MALDI-TOF/TOF mass spectrometry showed that differential quantitation using SILAC was highly reproducible (~8% coefficient of variation (CV)), and this variance was considerably lower than that achieved using automated 2-DE image analysis strategies (CV~25%). Both techniques revealed subtle alterations in cellular protein expression following uPA stimulation. However, due to the lower variances associated with the SILAC technique, smaller changes in expression of uPA-inducible proteins could be found with greater certainty.
- Description
- 8 page(s)
- Subject Keyword
- 030199 Analytical Chemistry not elsewhere classified
- Subject Keyword
- 030406 Proteins and Peptides
- Subject Keyword
- metabolic labeling
- Subject Keyword
- SILAC
- Subject Keyword
- two-dimensional gel electrophoresis
- Subject Keyword
- image analysis
- Subject Keyword
- MALDI-TOF/TOF
- Subject Keyword
- urokinase plasminogen activator
- Subject Keyword
- protein quantitation
- Resource Type
- journal article
- Organisation
- Macquarie University. Dept. of Chemistry and Biomolecular Sciences
- Organisation
- Macquarie University. Australian Proteome Analysis Facility (APAF)
- Organisation
- Macquarie University. Dept. of Statistics
- Identifier
- http://hdl.handle.net/1959.14/42511
- Identifier
- ISSN:1535-3907
- Identifier
- mq-rm-2007002065
- Language
- eng
- Reviewed
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