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Multiplexed Preparation of Biological Samples for Mass Spectrometry Using Gel Electrophoresis

  • Jeremy L. NorrisEmail author
  • Alan A. Doucette
Chapter

Abstract

In spite of the perception that gel electrophoresis has limited compatibility with modern mass spectrometry, polyacrylamide gel electrophoresis remain the gold standard for high resolution protein separation. Recent work has sought to capitalize on both the advantages of gel electrophoresis and mass spectrometry as a tool for protein separation and analysis, combining the two technologies into integrated workflows that have unique capabilities. This chapter discusses recent advances of this technology and applications for its use in mass spectrometry assays.

Keywords

Electrophoresis Fractionation Proteomics Quantitation Sample preparation 

References

  1. Beavis, R.C., and Chait, B.T. (1990). Rapid, sensitive analysis of protein mixtures by mass spectrometry. Proc Natl Acad Sci USA 87, 6873–6877.CrossRefGoogle Scholar
  2. Botelho, D., Wall, M.J., Vieira, D.B., Fitzsimmons, S., Liu, F., and Doucette, A. (2010). Top-down and bottom-up proteomics of SDS-containing solutions following mass-based separation. J Proteome Res 9, 2863–2870.CrossRefGoogle Scholar
  3. de Godoy, L.M., Olsen, J.V., Cox, J., Nielsen, M.L., Hubner, N.C., Frohlich, F., Walther, T.C., and Mann, M. (2008). Comprehensive mass-spectrometry-based proteome quantification of haploid versus diploid yeast. Nature 455, 1251–1254.CrossRefGoogle Scholar
  4. Fang, Y., Robinson, D.P., and Foster, L.J. (2010). Quantitative analysis of proteome coverage and recovery rates for upstream fractionation methods in proteomics. J Proteome Res 9, 1902–1912.CrossRefGoogle Scholar
  5. Gygi, S.P., Corthals, G.L., Zhang, Y., Rochon, Y., and Aebersold, R. (2000). Evaluation of two-dimensional gel electrophoresis-based proteome analysis technology. Proc Natl Acad Sci USA 97, 9390–9395.CrossRefGoogle Scholar
  6. Harkins, J.B.t., Katz, B.B., Pastor, S.J., Osucha, P., Hafeman, D.G., Witkowski, C.E., 2nd, and Norris, J.L. (2008). Parallel electrophoretic depletion, fractionation, concentration, and desalting of 96 complex biological samples for mass spectrometry. Anal Chem 80, 2734–2743.CrossRefGoogle Scholar
  7. Ikonomou, M.G., Blades, A.T., and Kebarle, P. (1990). Investigations of the electrospray interface for liquid chromatography mass spectrometry. Anal Chem 62, 957–967.CrossRefGoogle Scholar
  8. Lee, J.E., Kellie, J.F., Tran, J.C., Tipton, J.D., Catherman, A.D., Thomas, H.M., Ahlf, D.R., Durbin, K.R., Vellaichamy, A., Ntai, I., et al. (2009). A robust two-dimensional separation for top-down tandem mass spectrometry of the low-mass proteome. J Am Soc Mass Spectrom 20, 2183–2191.CrossRefGoogle Scholar
  9. Little, J.L., Wempe, M.F., and Buchanan, C.M. (2006). Liquid chromatography-mass spectrometry/mass spectrometry method development for drug metabolism studies: Examining lipid matrix ionization effects in plasma. J Chromatogr B Anal Technol Biomed Life Sci 833, 219–230.CrossRefGoogle Scholar
  10. Piersma, S.R., Fiedler, U., Span, S., Lingnau, A., Pham, T.V., Hoffmann, S., Kubbutat, M.H., and Jimenez, C.R. (2010). Workflow comparison for label-free, quantitative secretome proteomics for cancer biomarker discovery: method evaluation, differential analysis, and verification in serum. J Proteome Res 9, 1913–1922.CrossRefGoogle Scholar
  11. Puchades, M., Westman, A., Blennow, K., and Davidsson, P. (1999). Removal of sodium dodecyl sulfate from protein samples prior to matrix-assisted laser desorption/ionization mass spectrometry. Rapid Commun Mass Spectrom 13, 344–349.CrossRefGoogle Scholar
  12. Shevchenko, A., Tomas, H., Havlis, J., Olsen, J.V., and Mann, M. (2006). In-gel digestion for mass spectrometric characterization of proteins and proteomes. Nat Protoc 1, 2856–2860.CrossRefGoogle Scholar
  13. Shevchenko, A., Wilm, M., Vorm, O., and Mann, M. (1996). Mass spectrometric sequencing of proteins silver-stained polyacrylamide gels. Anal Chem 68, 850–858.CrossRefGoogle Scholar
  14. Tran, J.C. (2008). Development of Effective Proteomic Separations for Biological Analysis Using Mass Spectrometry. Thesis (Ph.D) – Dalhousie University, Halifax, Canada. In Chemistry (Halifax, NS, Dalhousie University), p. 177.Google Scholar
  15. Tran, J.C., and Doucette, A.A. (2008). Gel-eluted liquid fraction entrapment electrophoresis: An electrophoretic method for broad molecular weight range proteome separation. Anal Chem 80, 1568–1573.CrossRefGoogle Scholar
  16. Tran, J.C., and Doucette, A.A. (2009). Multiplexed Size Seperation of Intact Proteins in Solution Phase for Mass Spectrometry. Anal Chem 81, pp 6201–6209.Google Scholar
  17. Wessel, D., and Flugge, U.I. (1984). A method for the quantitative recovery of protein in dilute solution in the presence of detergents and lipids. Anal Biochem 138, 141–143.CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2011

Authors and Affiliations

  1. 1.Protein Discovery, Inc.KnoxvilleUSA
  2. 2.Dalhousie UniversityHalifaxCanada

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