Abstract

Phase I: The R43 phase of this project aims to develop and demonstrate a capillary gel electrophoresis (CGE) based multidimensional protein separation platform capable of providing significant analyte concentration and extremely high resolving power for handling complex protein mixtures prior to mass spectrometry detection. In comparison with conventional SDS-PAGE-based proteome approaches, the proposed CGE/capillary reversed-phase liquid chromatography (CRPLC) proteome technology will be highly automated and offer robust and high throughput fractionation of whole proteins for top-down proteomics while avoiding analyte dilution and loss in an integrated platform. By coupling with a nanoscale trypsin membrane reactor, the ultrafast proteolytic digestion of proteins resolved and eluted from the CGE-based separations enables the combined top-down/bottom-up characterization of post-translational modifications in complex proteomes. Research efforts in the R43 phase will address early implementation, optimization, and demonstration of the proposed proteome technology using yeast cell lysates.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Small Business Innovation Research Grants (SBIR) - Phase II (R44)
Project #
5R44CA107988-03
Application #
7103624
Study Section
Special Emphasis Panel (ZCA1-SRRB-C (J1))
Program Officer
Heath, Anne K
Project Start
2004-04-07
Project End
2008-07-31
Budget Start
2006-08-15
Budget End
2007-07-31
Support Year
3
Fiscal Year
2006
Total Cost
$816,936
Indirect Cost

  Institution

Name
Calibrant Biosystems, Inc.
Department
Type
DUNS #
616684036
City
Gaithersburg
State
MD
Country
United States
Zip Code
20878

  Publications

Lee, Cheng S; Balgley, Brian M (2009) Capillary isoelectric focusing/reversed phase liquid chromatography/mass spectrometry. Methods Mol Biol 492:233-40
Balgley, Brian M; Wang, Weijie; Song, Tao et al. (2008) Evaluation of confidence and reproducibility in quantitative proteomics performed by a capillary isoelectric focusing-based proteomic platform coupled with a spectral counting approach. Electrophoresis 29:3047-54
Balgley, Brian M; Laudeman, Tom; Yang, Li et al. (2007) Comparative evaluation of tandem MS search algorithms using a target-decoy search strategy. Mol Cell Proteomics 6:1599-608
Fang, Xueping; Yang, Li; Wang, Weijie et al. (2007) Comparison of electrokinetics-based multidimensional separations coupled with electrospray ionization-tandem mass spectrometry for characterization of human salivary proteins. Anal Chem 79:5785-92
Wang, Weijie; Guo, Tong; Song, Tao et al. (2007) Comprehensive yeast proteome analysis using a capillary isoelectric focusing-based multidimensional separation platform coupled with ESI-MS/MS. Proteomics 7:1178-87
Wang, Weijie; Guo, Tong; Rudnick, Paul A et al. (2007) Membrane proteome analysis of microdissected ovarian tumor tissues using capillary isoelectric focusing/reversed-phase liquid chromatography-tandem MS. Anal Chem 79:1002-9
Guo, Tong; Wang, Weijie; Rudnick, Paul A et al. (2007) Proteome analysis of microdissected formalin-fixed and paraffin-embedded tissue specimens. J Histochem Cytochem 55:763-72
Guo, Tong; Rudnick, Paul A; Wang, Weijie et al. (2006) Characterization of the human salivary proteome by capillary isoelectric focusing/nanoreversed-phase liquid chromatography coupled with ESI-tandem MS. J Proteome Res 5:1469-78
Guo, Tong; Lee, Cheng S; Wang, Weijie et al. (2006) Capillary separations enabling tissue proteomics-based biomarker discovery. Electrophoresis 27:3523-32
Wang, Yueju; Balgley, Brian M; Rudnick, Paul A et al. (2005) Effects of chromatography conditions on intact protein separations for top-down proteomics. J Chromatogr A 1073:35-41

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