Quantitative 2-D gel electrophoresis-based expression proteomics of albumin and IgG immunodepleted plasma

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DOIResolve DOI: http://doi.org/10.1016/j.jchromb.2008.01.052
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TypeArticle
Journal titleJournal of Chromatography B
Volume865
Issue1-2
Pages147152; # of pages: 6
SubjectPlasma proteome; Albumin; IgG; Immunodepletion; ELISA; 2-DGE software; Expression proteomics
AbstractProteomic analysis of plasma is challenging because of its large dynamic range, which prevents the detection of low abundance proteins. Immunodepletion of high abundance proteins, such as albumin and IgG, has emerged as a favored technology to overcome this problem; however its suitability in quantitative expression proteomics has not yet been adequately addressed. In this study, albumin and IgG immunodepletion was evaluated by ELISAs and the reproducibility of depletion was tested with 2-DGE. Depletion of plasma resulted in removal of 62 ± 1.2% of the total protein, 93 ± 1.4% of the albumin (0.43 μg/μL, residual), and 94 ± 1.5% of the IgG (0.21 μg/μL, residual). These results were confirmed by immunoblotting. Computerized image analysis of 2-D gels using Progenesis SameSpots software revealed an enhancement in the number of visible spots (675–1325), with 10 ± 6% inter-gel variability in spot density. LC–ESI-MS/MS identification of newly resolved protein spots further validated the procedure. An innovative application of the software employed led to identification of 11 proteins lost non-specifically during depletion. This study demonstrates the effectiveness of immunodepletion of albumin and IgG in quantitative 2-DGE-based differential analysis of plasma proteins.
Publication date
AffiliationNRC Institute for Marine Biosciences; National Research Council Canada
Peer reviewedYes
NRC number1768
NPARC number3538331
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Record identifierbe9df090-02fa-4892-b1ca-a45d1f190639
Record created2009-03-01
Record modified2016-05-09
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