The proteomic reactor facilitates the analysis of affinity-purified proteins by mass spectrometry: application for identifying ubiquitinated proteins in human cells

J Proteome Res. 2007 Jan;6(1):298-305. doi: 10.1021/pr060438j.


Mass spectrometry (MS) coupled to affinity purification is a powerful approach for identifying protein-protein interactions and for mapping post-translational modifications. Prior to MS analysis, affinity-purified proteins are typically separated by gel electrophoresis, visualized with a protein stain, excised, and subjected to in-gel digestion. An inherent limitation of this series of steps is the loss of protein sample that occurs during gel processing. Although methods employing in-solution digestion have been reported, they generally suffer from poor reaction kinetics. In the present study, we demonstrate an application of a microfluidic processing device, termed the Proteomic Reactor, for enzymatic digestion of affinity-purified proteins for liquid chromatography tandem mass spectrometry (LC-MS/MS) analysis. Use of the Proteomic Reactor enabled the identification of numerous ubiquitinated proteins in a human cell line expressing reduced amounts of the ubiquitin-dependent chaperone, valosin-containing protein (VCP). The Proteomic Reactor is a novel technology that facilitates the analysis of affinity-purified proteins and has the potential to aid future biological studies.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine Triphosphatases / chemistry
  • Cell Cycle Proteins / chemistry
  • Cell Line
  • Cell Line, Tumor
  • Chromatography, Liquid
  • Humans
  • Kinetics
  • Mass Spectrometry / methods*
  • Microfluidic Analytical Techniques
  • Plasmids / metabolism
  • Proteasome Endopeptidase Complex / metabolism
  • Proteome
  • Proteomics / instrumentation*
  • Proteomics / methods*
  • Ubiquitin / chemistry*
  • Valosin Containing Protein


  • Cell Cycle Proteins
  • Proteome
  • Ubiquitin
  • Proteasome Endopeptidase Complex
  • Adenosine Triphosphatases
  • VCP protein, human
  • Valosin Containing Protein