HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) Supplemental Data All Versions of this Article: M700132-MCP200v1 M700132-MCP200v2 6/10/1809    most recent Submit a response Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Glossary Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Stahl-Zeng, J. Articles by Domon, B. Search for Related Content PubMed PubMed Citation Articles by Stahl-Zeng, J. Articles by Domon, B. Social Bookmarking                      What's this?

Molecular & Cellular Proteomics 6:1809-1817, 2007.
© 2007 by The American Society for Biochemistry and Molecular Biology, Inc.

---

Research

High Sensitivity Detection of Plasma Proteins by Multiple Reaction Monitoring of N-Glycosites^*,S

Jianru Stahl-Zeng^,, Vinzenz Lange^,¶,||, Reto Ossola^¶, Katrin Eckhardt^||, Wilhelm Krek^||, Ruedi Aebersold^¶,**, and Bruno Domon^¶,

From Applied Biosystems, 64293 Darmstadt, Germany, ^¶ Institute of Molecular Systems Biology and ^|| Competence Center for Systems Physiology and Metabolic Diseases, Institute of Cell Biology/ETH Zurich, CH-8093 Zurich, Switzerland, ^** Faculty of Sciences, University of Zurich, CH-8006 Zurich, Switzerland, and Institute for Systems Biology, Seattle, Washington 98103

The detection and quantification of plasma (serum) proteins at or below the ng/ml concentration range are of critical importance for the discovery and evaluation of new protein biomarkers. This has been achieved either by the development of high sensitivity ELISA or other immunoassays for specific proteins or by the extensive fractionation of the plasma proteome followed by the mass spectrometric analysis of the resulting fractions. The first approach is limited by the high cost and time investment for assay development and the requirement of a validated target. The second, although reasonably comprehensive and unbiased, is limited by sample throughput. Here we describe a method for the detection of plasma proteins at concentrations in the ng/ml or sub-ng/ml range and their accurate quantification over 5 orders of magnitude. The method is based on the selective isolation of N-glycosites from the plasma proteome and the detection and quantification of targeted peptides in a quadrupole linear ion trap instrument operated in the multiple reaction monitoring (MRM) mode. The unprecedented sensitivity of the mass spectrometric analysis of minimally fractionated plasma samples is the result of the significantly reduced sample complexity of the isolated N-glycosites compared with whole plasma proteome digests and the selectivity of the MRM process. Precise quantification was achieved via stable isotope dilution by adding ¹³C- and/or ¹⁵N-labeled reference analytes. We also demonstrate the possibility of significantly expanding the number of MRM measurements during one single LC-MS run without compromising sensitivity by including elution time constraints for the targeted transitions, thus allowing quantification of large sets of peptides in a single analysis.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				N. L. Anderson, N. G. Anderson, T. W. Pearson, C. H. Borchers, A. G. Paulovich, S. D. Patterson, M. Gillette, R. Aebersold, and S. A. Carr A Human Proteome Detection and Quantitation Project Mol. Cell. Proteomics, May 1, 2009; 8(5): 883 - 886. [Abstract] [Full Text] [PDF]

				J. A. Mead, L. Bianco, V. Ottone, C. Barton, R. G. Kay, K. S. Lilley, N. J. Bond, and C. Bessant MRMaid, the Web-based Tool for Designing Multiple Reaction Monitoring (MRM) Transitions Mol. Cell. Proteomics, April 1, 2009; 8(4): 696 - 705. [Abstract] [Full Text] [PDF]

				A. K. Yocum and A. M. Chinnaiyan Current affairs in quantitative targeted proteomics: multiple reaction monitoring-mass spectrometry Brief Funct Genomic Proteomic, March 11, 2009; (2009) eln056v1. [Abstract] [Full Text] [PDF]

				K. Helsens, E. Timmerman, J. Vandekerckhove, K. Gevaert, and L. Martens Peptizer, a Tool for Assessing False Positive Peptide Identifications and Manually Validating Selected Results Mol. Cell. Proteomics, December 1, 2008; 7(12): 2364 - 2372. [Abstract] [Full Text] [PDF]

				W.-J. Qian, D. T. Kaleta, B. O. Petritis, H. Jiang, T. Liu, X. Zhang, H. M. Mottaz, S. M. Varnum, D. G. Camp II, L. Huang, et al. Enhanced Detection of Low Abundance Human Plasma Proteins Using a Tandem IgY12-SuperMix Immunoaffinity Separation Strategy Mol. Cell. Proteomics, October 1, 2008; 7(10): 1963 - 1973. [Abstract] [Full Text] [PDF]

				V. Lange, J. A. Malmstrom, J. Didion, N. L. King, B. P. Johansson, J. Schafer, J. Rameseder, C.-H. Wong, E. W. Deutsch, M.-Y. Brusniak, et al. Targeted Quantitative Analysis of Streptococcus pyogenes Virulence Factors by Multiple Reaction Monitoring Mol. Cell. Proteomics, August 1, 2008; 7(8): 1489 - 1500. [Abstract] [Full Text] [PDF]

				A. J. Hulsmeier, P. Paesold-Burda, and T. Hennet N-Glycosylation Site Occupancy in Serum Glycoproteins Using Multiple Reaction Monitoring Liquid Chromatography-Mass Spectrometry Mol. Cell. Proteomics, December 1, 2007; 6(12): 2132 - 2138. [Abstract] [Full Text] [PDF]

				H. Keshishian, T. Addona, M. Burgess, E. Kuhn, and S. A. Carr Quantitative, Multiplexed Assays for Low Abundance Proteins in Plasma by Targeted Mass Spectrometry and Stable Isotope Dilution Mol. Cell. Proteomics, December 1, 2007; 6(12): 2212 - 2229. [Abstract] [Full Text] [PDF]