One step microelectroelution concentration method for efficient coupling of sodium dodecylsulfate gel electrophoresis and matrix-assisted laser desorption time-of-flight mass spectrometry for protein analysis

  • Nigel J. Clarke
  • Feng Li
  • Andy J. Tomlinson
  • Stephen NaylorEmail author
Short Communication


The coupling of the widely used separation technique of conventional sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE) with the mass accuracy measurement capability of mass Spectrometry (MS) provides a very powerful analytical technique. However, at present, there is no simple, definitive method for coupling the two methods. Typically, separated proteins are extracted from the gel, either as the native protein or as a peptide mixture after in-gel proteolytic digestion, and then analyzed by mass Spectrometry. However, the various extraction techniques described previously have been labor intensive and require a large number of steps. The mass Spectrometry analysis of very low concentrations of in vivo derived proteins requires minimum sample handling and on-line concentration. Therefore, we have developed an efficient microelectroelution technique that is applied in a single step manner and contains an on-line concentration device. Initial results from this system have shown a high efficiency of analyte elution from the gel and a simple, robust technique for the coupling of SDS-containing gels with MALDI-TOF-MS analysis and a capability of analyzing proteins at the subpicomole level.


Carbonic Anhydrase Mass Accuracy Fuse Silica Capillary Protein Mixture Mass Accuracy Measurement 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. 1.
    Clauser, K. R.; Hall, S. C.; Smith, D. M.; Webb, J. W.; Andrews, L. E.; Tran, H. M.; Epstein, L. B.; Burlingame, A. L. Proc. Natl. Acad. Sci. USA 1995, 92, 5072.CrossRefGoogle Scholar
  2. 2.
    Patterson, S. D.; Aebersold, R. Electrophoresis 1995, 16, 1791.CrossRefGoogle Scholar
  3. 3.
    Houthaeve, T.; Gausepohl, H.; Mann, M.; Ashman, K. FEBS Lett. 1995, 376, 91.CrossRefGoogle Scholar
  4. 4.
    Mortz, E.; Vorm, O.; Mann, M.; Roepstroff, P. Biol Mass Spectrom. 1994, 23, 249.CrossRefGoogle Scholar
  5. 5.
    Eckerskorn, C.; Strupat, K.; Karas, M.; Hillenkamp, F. Ekctrophoresis 1992, 13, 664.CrossRefGoogle Scholar
  6. 6.
    Vestling, M. M.; Feneslau, C. Anal. Chem. 1994, 66, 471.CrossRefGoogle Scholar
  7. 7.
    Rasmussen, H. H.; Mortz, E.; Mann, M.; Roepstroff, P.; Cells, J. E. Electrophoresis 1994, 15, 406.CrossRefGoogle Scholar
  8. 8.
    Habel, S.; Jensen, C.; Anderson, S. O.; Roepstorff, P. Protein Sci. 1995, 4, 394.CrossRefGoogle Scholar
  9. 9.
    Schumacher, M.; Glocker, M. O.; Wunderlin, M.; Przybylski, M. Electrophoresis 1996, 17, 848.CrossRefGoogle Scholar
  10. 10.
    Zhang, W. Z.; Czernik, A. J.; Yungwirth, T.; Aebersold, R.; Chait, B. T. Prot. Sci. 1994, 3, 677.CrossRefGoogle Scholar
  11. 11.
    Shevchenko, A.; Wilm, M.; Vorm, O.; Mann, M. Anal. Chem. 1996, 68, 850.CrossRefGoogle Scholar
  12. 12.
    Bai, J.; Qian, M. G.; Liu, Y.; Liang, X.; Lubman, D. M. Anal. Chem. 1995, 67, 1705.CrossRefGoogle Scholar
  13. 13.
    Loo, R. R. O.; Stevenson, T. I.; Mitchell, C.; Loo, J. A.; Andrews, P. C. Anal. Chem. 1996, 68, 1910.CrossRefGoogle Scholar
  14. 14.
    Loo, R. R. O.; Mitchell, C.; Stevenson, T. I.; Martin, S. A.; Hines, W. M.; Juhasz, P.; Patterson, D. H.; Pelteir, J. M.; Loo, J. A.; Andrews, P. C. Electrophoresis 1997, 18, 382.CrossRefGoogle Scholar
  15. 15.
    Tomlinson, A. J.; Benson, L. M.; Braddock, W. D.; Oda, R. P.; Naylor, S. J. High Resolut. Chromatogr. 1995, 18, 381.CrossRefGoogle Scholar
  16. 16.
    Tomlinson, A. J.; Jameson, S.; Naylor, S. J. Chromatogr. A 1996, 744, 273.CrossRefGoogle Scholar
  17. 17.
    Tomlinson, A. J.; Benson, L. M.; Jameson, S.; Johnson, D. H.; Naylor, S. J. Am. Soc. Mass Spectrom. 1997, 8, 15.CrossRefGoogle Scholar
  18. 18.
    Hare, P. E. Methods Enzymol. 1977, 43, 3.CrossRefGoogle Scholar
  19. 19.
    Pharmacia LKB Technology AB, Separation File 110, PhastsystemTM, Uppsala Sweden.Google Scholar
  20. 20.
    Loo, R. R. O.; Dales, N.; Andrews, P. C. Prot. Sci. 1994, 3, 1975.CrossRefGoogle Scholar
  21. 21.
    Tuszynski, G. P.; Warren, L. Anal. Biochem. 1975, 67, 55.CrossRefGoogle Scholar
  22. 22.
    Stearne, P. A.; van Driel, I. R.; Grego, B.; Simpson, R.; Goding, J. W. J. Immunol. 1985, 109, 76.Google Scholar
  23. 23.
    Konigsberg, W. H.; Henderson, L. Meth. Enzymol. 1983, 91, 254.CrossRefGoogle Scholar

Copyright information

© American Society for Mass Spectrometry 1998

Authors and Affiliations

  • Nigel J. Clarke
    • 1
  • Feng Li
    • 1
  • Andy J. Tomlinson
    • 1
  • Stephen Naylor
    • 1
    Email author
  1. 1.Biomedical Mass Spectrometry FacilityDepartment of Biochemistry and Molecular Biology and Department of Pharmacology and Clinical Pharmacology, Mayo ClinicRochesterUSA

Personalised recommendations