Antibodies and Activity Measurements for the Detection of O-GlcNAc Transferase and Assay of its Substrate, UDP-GlcNAc

  • Tony Lefebvre
  • Ludivine Drougat
  • Stephanie Olivier-Van Stichelen
  • Jean-Claude Michalski
  • Anne-Sophie Vercoutter-Edouart
Part of the Methods in Molecular Biology book series (MIMB, volume 1022)


Since the discovery of O-GlcNAc modification (O-GlcNAcylation) 20 years ago, much attention has been given to OGT (O-GlcNAc transferase), the unique enzyme responsible for the nuclear and cytosolic O-GlcNAcylation processes. This review focuses on protocols that are routinely used to analyze OGT expression and activity. First are detailed techniques using rabbit polyclonal anti-OGT antibodies, namely, Western blot, (co-)immunoprecipitation, and immunofluorescence. We also describe the measurement of OGT activity by using synthetic peptides as acceptors and radiolabeled UDP-GlcNAc. Finally, a sensitive HPAEC-based technique to measure the cellular content of UDP-GlcNAc, the donor substrate of OGT, is described in detail.

Key words

O-GlcNAc transferase Polyclonal anti-OGT antibodies OGT activity assay UDP-GlcNAc content assay 


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Copyright information

© Springer Science+Business Media New York 2013

Authors and Affiliations

  • Tony Lefebvre
    • 1
  • Ludivine Drougat
    • 1
  • Stephanie Olivier-Van Stichelen
    • 1
  • Jean-Claude Michalski
    • 1
  • Anne-Sophie Vercoutter-Edouart
    • 1
  1. 1.Unit of Structural and Functional GlycobiologyUniversity of Lille 1Villeneuve d’AscqFrance

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