In-Depth Analysis of Protein Phosphorylation by Multidimensional Ion Exchange Chromatography and Mass Spectrometry

  • Maria P. Alcolea
  • Pedro R. Cutillas
Part of the Methods in Molecular Biology book series (MIMB, volume 658)


Protein phosphorylation controls fundamental biological functions that are often deregulated in disease. Therefore, system-level understanding of complex pathophysiological processes requires methods that can be used to profile and quantify protein phosphorylation as comprehensively as possible. Here we present a detailed protocol to enrich phosphopeptides from total cell lysates in a form amenable to downstream analysis by mass spectrometry. Using these techniques, we have detected several thousands of phosphorylation sites in the NIH-3T3 cell line.

Key words

Phosphoproteomics cell signaling cancer quantification 


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Copyright information

© Springer Science+Business Media, LLC 2010

Authors and Affiliations

  • Maria P. Alcolea
    • 1
  • Pedro R. Cutillas
    • 1
  1. 1.Analytical Signalling Group, Centre for Cell SignallingInstitute of Cancer, Bart’s and the London School of Medicine, Queen Mary University of LondonLondonUK

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