Improving image analysis in 2DGE-based redox proteomics by labeling protein carbonyl with fluorescent hydroxylamine |
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Authors: | H Fai Poon Laila Abdullah Jon Reed Sarah M Doore Cyndi Laird Venkat Mathura Michael Mullan Fiona Crawford |
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Institution: | (1) Roskamp Institute, 2040 Whitfield Ave., 34243 Sarasota, FL, USA |
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Abstract: | Recent advances in redox proteomics have provided significant insight into the role of oxidative modifications in cellular
signalling and metabolism. At present, these techniques rely heavily on Western blots to visualize the oxidative modification
and corresponding two dimensional (2D) gels for detection of total protein levels, resulting in the duplication of efforts.
A major limitation associated with this methodology includes problematic matching up of gels and blots due to the differences
in processing and/or image acquisition. In this study, we present a new method which allows detection of protein oxidation
and total protein on the same gel to improve matching in image analysis. Furthermore, the digested protein spots are compatible
with standard MALDI mass spectrometry protein identification. The methodology highlighted here may be useful in facilitating
the development of biomarkers, assessing potential therapeutic targets and elucidating new mechanisms of redox signalling
in redox-related conditions. |
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Keywords: | Indexing terms" target="_blank">Indexing terms electrophoresis gel two-dimensional oxidation-reduction protein processing post-translational |
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