Quantitative mass spectrometry by isotope dilution and multiple reaction monitoring (MRM)

Paul Russo*, Brian L. Hood, Nicholas W. Bateman, Thomas P. Conrads

*Corresponding author for this work

Research output: Chapter in Book/Report/Conference proceedingChapterpeer-review

11 Scopus citations


Selected reaction monitoring (SRM) is used in molecular profiling to detect and quantify specific known proteins in complex mixtures. Using isotope dilution (Barnidge et al., Anal Chem 75(3):445–451, 2003) methodologies, peptides can be quantified without the need for an antibody-based method. Selected reaction monitoring assays employ electrospray ionization mass spectrometry (ESI-MS) followed by two stages of mass selection: a first stage where the mass of the peptide ion is selected and, after fragmentation by collision-induced dissociation (CID), a second stage (tandem MS) where either a single (e.g., SRM) or multiple (multiple reaction monitoring, MRM) specific peptide fragment ions are transmitted for detection. The MRM experiment is accomplished by specifying the parent masses of the selected endogenous and isotope-labeled peptides for MS/MS fragmentation and then monitoring fragment ions of interest, using their intensities/abundances and relative ratios to quantify the parent protein of interest. In this example protocol, we will utilize isotope dilution MRM-MS to quantify in absolute terms the total levels of the protein of interest, ataxia telangiectasia mutated (ATM) serine/threonine protein kinase. Ataxia telangiectasia mutated (ATM) phosphorylates several key proteins that initiate activation of the DNA damage checkpoint leading to cell cycle arrest.

Original languageEnglish
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc.
Number of pages20
StatePublished - 2017
Externally publishedYes

Publication series

NameMethods in Molecular Biology
ISSN (Print)1064-3745


  • Ataxia telangiectasia mutated
  • Collision-induced dissociation
  • Isotope dilution
  • Mass spectrometry
  • Multiple reaction monitoring
  • Peptide
  • Protein
  • Proteomics
  • Selected reaction monitoring


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