NP-40 reduces contamination by endogenous biotinylated carboxylases during purification of biotin tagged nuclear proteins

Protein Expr Purif. 2013 May;89(1):80-3. doi: 10.1016/j.pep.2013.02.015. Epub 2013 Mar 7.


We describe here a simple procedure for greatly reducing contamination of nuclear extracts by naturally biotinylated cytoplasmic carboxylases, which represent a major source of non-specific background when employing BirA-mediated biotinylation tagging for the purification and characterization of nuclear protein complexes by mass spectrometry. We show that the use of 0.5% of the non-ionic detergent Nonidet-40 (NP-40) during cell lysis and nuclei isolation is sufficient to practically eliminate contamination of nuclear extracts by carboxylases and to greatly reduce background signals in downstream mass spectrometric analyses.

Publication types

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

MeSH terms

  • Biotin / chemistry*
  • Biotinylation
  • Carbon-Nitrogen Ligases / chemistry
  • Cell Extracts / chemistry
  • Cell Extracts / isolation & purification
  • Cell Nucleus / chemistry
  • Escherichia coli Proteins / chemistry
  • Nuclear Proteins / isolation & purification*
  • Octoxynol
  • Polyethylene Glycols / chemistry*
  • Repressor Proteins / chemistry


  • Cell Extracts
  • Escherichia coli Proteins
  • Nuclear Proteins
  • Repressor Proteins
  • Polyethylene Glycols
  • Biotin
  • Octoxynol
  • Nonidet P-40
  • Carbon-Nitrogen Ligases
  • birA protein, E coli