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

@article{citeulike:2803511, abstract = {Proteomic 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 microg/microL, residual), and 94+/-1.5\% of the IgG (0.21 microg/microL, 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.}, address = {Department of Pediatrics, University of Western Ontario, London, ON, Canada.}, author = {Seferovic, M. D. and Krughkov, V. and Pinto, D. and Han, V. K. and Gupta, M. B. }, citeulike-article-id = {2803511}, doi = {10.1016/j.jchromb.2008.01.052}, issn = {1570-0232}, journal = {Journal of chromatography. B, Analytical technologies in the biomedical and life sciences}, month = {April}, number = {1-2}, pages = {147--152}, posted-at = {2008-05-16 02:58:23}, priority = {2}, title = {Quantitative 2-D gel electrophoresis-based expression proteomics of albumin and IgG immunodepleted plasma.}, url = {http://dx.doi.org/10.1016/j.jchromb.2008.01.052}, volume = {865}, year = {2008} }

TY - JOUR ID - citeulike:2803511 L3 - citeulike-article-id:2803511 TI - Quantitative 2-D gel electrophoresis-based expression proteomics of albumin and IgG immunodepleted plasma. JF - Journal of chromatography. B, Analytical technologies in the biomedical and life sciences VL - 865 IS - 1-2 SP - 147 EP - 152 AD - Department of Pediatrics, University of Western Ontario, London, ON, Canada. SN - 1570-0232 N2 - Proteomic 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 microg/microL, residual), and 94+/-1.5% of the IgG (0.21 microg/microL, 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. AU - Seferovic, MD AU - Krughkov, V AU - Pinto, D AU - Han, VK AU - Gupta, MB PY - 2008/04/01/ UR - http://dx.doi.org/10.1016/j.jchromb.2008.01.052 ER -