Systematic comparison of label-free, metabolic labeling, and isobaric chemical labeling for quantitative proteomics on LTQ Orbitrap Velos

J Proteome Res. 2012 Mar 2;11(3):1582-90. doi: 10.1021/pr200748h. Epub 2012 Feb 16.


A variety of quantitative proteomics methods have been developed, including label-free, metabolic labeling, and isobaric chemical labeling using iTRAQ or TMT. Here, these methods were compared in terms of the depth of proteome coverage, quantification accuracy, precision, and reproducibility using a high-performance hybrid mass spectrometer, LTQ Orbitrap Velos. Our results show that (1) the spectral counting method provides the deepest proteome coverage for identification, but its quantification performance is worse than labeling-based approaches, especially the quantification reproducibility; (2) metabolic labeling and isobaric chemical labeling are capable of accurate, precise, and reproducible quantification and provide deep proteome coverage for quantification; isobaric chemical labeling surpasses metabolic labeling in terms of quantification precision and reproducibility; and (3) iTRAQ and TMT perform similarly in all aspects compared in the current study using a CID-HCD dual scan configuration. On the basis of the unique advantages of each method, we provide guidance for selection of the appropriate method for a quantitative proteomics study.

Publication types

  • Comparative Study
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Algorithms
  • Bacterial Proteins / chemistry
  • Bacterial Proteins / metabolism*
  • Isotope Labeling
  • Peptide Fragments / chemistry
  • Peptide Mapping / methods
  • Proteome / chemistry
  • Proteome / metabolism*
  • Proteomics
  • Pseudomonas putida / metabolism
  • Reproducibility of Results
  • Staining and Labeling / methods*
  • Tandem Mass Spectrometry / methods*


  • Bacterial Proteins
  • Peptide Fragments
  • Proteome