Antibodies to biotin enable large-scale detection of biotinylation sites on proteins

Nat Methods. 2017 Dec;14(12):1167-1170. doi: 10.1038/nmeth.4465. Epub 2017 Oct 16.


Although purification of biotinylated molecules is highly efficient, identifying specific sites of biotinylation remains challenging. We show that anti-biotin antibodies enable unprecedented enrichment of biotinylated peptides from complex peptide mixtures. Live-cell proximity labeling using APEX peroxidase followed by anti-biotin enrichment and mass spectrometry yielded over 1,600 biotinylation sites on hundreds of proteins, an increase of more than 30-fold in the number of biotinylation sites identified compared to streptavidin-based enrichment of proteins.

MeSH terms

  • Antibodies / metabolism*
  • Biotechnology / methods
  • Biotin / metabolism*
  • Biotinylation
  • Chromatography, Liquid
  • HEK293 Cells
  • Humans
  • Jurkat Cells
  • Peptides / chemistry*
  • Proteins / chemistry*
  • Proteins / isolation & purification
  • Staining and Labeling
  • Streptavidin / metabolism
  • Tandem Mass Spectrometry


  • Antibodies
  • Peptides
  • Proteins
  • Biotin
  • Streptavidin