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Protein identification from two-dimensional gel electrophoresis by connecting ZipTip with a home-assembled nanoflow system

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Abstract

Mass spectrometry (MS) is a powerful technique for protein identification in proteomic research. Two-dimensional gel electrophoresis (2-DE) combined with MS is a significant method for protein separation and identification. For protein identification, peptide sequencing is usually carried out by an effective but expensive nano-flow liquid chromatographic system combined to tandem mass spectrometry (MS/MS). However, protein identification based on such method is time-consuming, and contamination may occur as a result of column overloading. In this study, we establish an alternative nanoscale system for protein identification using MS/MS. The system consists of several devices that can be purchased from commercial sources and can be connected to an electrospray ionization quadrupole-time of flight (ESI-Q-TOF) MS in order to analyze proteins from 2D gels. This inexpensive strategy provides an attractive alternative method for rapid identification of proteins using a nanospray source. In addition, the device is disposable so that contamination is avoided. It is shown that peptide sequencing based on this device using ESI-Q-TOF MS is accomplished within 10 min.

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References

  1. Nomura F, Tomonaga T, Sogawa K, Wu D, Ohashi T (2007) Application of proteomic technologies to discover and identify biomarkers for excessive alcohol consumption: a review. J Chromatogr B 855:35–41

    Article  CAS  Google Scholar 

  2. Ortea PM, Allis O, Healy BM, Lehane M, Shuilleabháin AA, Furey A, James KJ (2004) Determination of toxic cyclic heptapeptides by liquid chromatography with detection using ultra-violet, protein phosphatase assay and tandem mass spectrometry. Chemosphere 55:1395–1402

    Article  CAS  Google Scholar 

  3. Chalmers MJ, Gaskell SJ (2000) Advances in mass spectrometry for proteome analysis. Curr Opin Biotechnol 11:384–390

    Article  CAS  Google Scholar 

  4. Mo W, Karger BL (2002) Analytical aspects of mass spectrometry and proteomics. Curr Opin Chem Biol 6:666–675

    Article  CAS  Google Scholar 

  5. Lin D, Tabb DL, Yates JR III (2003) Large-scale protein identification using mass spectrometry. Biochim Biophys Acta 1646:1–10

    CAS  Google Scholar 

  6. Griffin TJ, Aebersold R (2001) Advances in proteome analysis by mass spectrometry. J Biol Chem 276:45497–45500

    Article  CAS  Google Scholar 

  7. Görg A, Obermaier C, Boguth G, Harder A, Scheibe B, Wildgruber R, Weiss W (2000) The current state of two-dimensional electrophoresis with immobilized pH gradients. Electrophoresis 21:1037–1053

    Article  Google Scholar 

  8. Washburn MP, Yates JR III (2000) Analysis of the microbial proteome. Curr Opin Microbiol 3:292–297

    Article  CAS  Google Scholar 

  9. Hanash SM (2000) Biomedical applications of two-dimensional electrophoresis using immobilized pH gradients: current status. Electrophoresis 21:1202–1209

    Article  CAS  Google Scholar 

  10. Rabilloud T (2002) Two-dimensional gel electrophoresis in proteomics: old, old fashioned, but it still climbs up the mountains. Proteomics 2:3–10

    Article  CAS  Google Scholar 

  11. Graves PR, Haystead TAJ (2002) Molecular biologist’s guide to proteomics. Microbiol Mol Biol Rev 66:39–63

    Article  CAS  Google Scholar 

  12. Patterson SD, Aebersold RH (2003) Proteomics: the first decade and beyond. Nat Genet 33:311–323

    Article  CAS  Google Scholar 

  13. MacCoss MJ, Yates JR III (2001) Proteomics: analytical tools and techniques. Curr Opin Clin Nutr Metab Care 4:369–375

    Article  CAS  Google Scholar 

  14. http://www.millipore.com/

  15. Chen YY, Lin SY, Yeh YY, Hsiao HH, Wu CY, Chen ST, Wang AHJ (2005) A modified protein precipitation procedure for efficient removal of albumin from serum. Electrophoresis 26:2117–2127

    Article  CAS  Google Scholar 

  16. Williams JG, Tomer KB (2004) Disposable chromatography for a high-throughput nano-ESI/MS and nano-ESI/MS-MS platform. J Am Soc Mass Spectrom 15:1333–1340

    Article  CAS  Google Scholar 

  17. Lu CY, Wu CY, Lin CH (2007) Protein identification by syringe pump-driven reversed-phase LC-MS/MS. Anal Biochem 368:123–129

    Article  CAS  Google Scholar 

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Acknowledgements

The authors are grateful to the National Science Council (NSC 98-2113-M-037-010 and 97-2113-M-037-008-MY2), Kaohsiung Medical University (Q-097021 and Q-098004) and Center of Excellence for Environmental Medicine (Kaohsiung Medical University) for financial support of this work. The authors are also grateful to the Center for Resources, Research and Development (Kaohsiung Medical University) for instrumental support of this work.

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Authors and Affiliations

  1. Department of Fragrance and Cosmetic Science, College of Pharmacy, Kaohsiung Medical University, Kaohsiung, 807, Taiwan

    Chia-Hsien Feng

  2. Center for Resources, Research and Development/Graduate Institute of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, 807, Taiwan

    Chi-Hsien Chou

  3. Department of Biochemistry, College of Medicine/Center of Excellence for Environmental Medicine, Kaohsiung Medical University, Kaohsiung, 807, Taiwan

    Lea-Yea Chuang & Chi-Yu Lu

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  1. Chia-Hsien Feng
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  2. Chi-Hsien Chou
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Correspondence to Chi-Yu Lu.

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Feng, CH., Chou, CH., Chuang, LY. et al. Protein identification from two-dimensional gel electrophoresis by connecting ZipTip with a home-assembled nanoflow system. Microchim Acta 170, 179–185 (2010). https://doi.org/10.1007/s00604-010-0399-9

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  • DOI: https://doi.org/10.1007/s00604-010-0399-9

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