Using sequential immunoprecipitation and mass spectrometry to identify methylation of NF-κB

Methods Mol Biol. 2015;1280:383-93. doi: 10.1007/978-1-4939-2422-6_23.


Posttranslational modifications have long been known to play an essential role in the regulation of NF-κB activity. In the past few years, in addition to more traditional modifications such as phosphorylation, the p65 subunit of NF-κB has been found to be methylated at multiple sites. Here, we describe procedures for using immunoprecipitation and mass spectrometry to identify the methylation sites of p65.

Publication types

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

MeSH terms

  • Cell Culture Techniques
  • Cell Line
  • Humans
  • Immunoprecipitation* / methods
  • In Vitro Techniques
  • Interleukin-1beta / pharmacology
  • Mass Spectrometry* / methods
  • Methylation / drug effects
  • NF-kappa B / metabolism*
  • Phosphorylation
  • Transcription Factor RelA / metabolism


  • Interleukin-1beta
  • NF-kappa B
  • Transcription Factor RelA