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Mol Cell Proteomics DOI:10.1074/mcp.M113.036095

Targeted peptide measurements in biology and medicine: best practices for mass spectrometry-based assay development using a fit-for-purpose approach.

Publication TypeJournal Article
Year of Publication2014
AuthorsCarr, SA, Abbatiello, SE, Ackermann, BL, Borchers, C, Domon, B, Deutsch, EW, Grant, RP, Hoofnagle, AN, Hüttenhain, R, Koomen, JM, Liebler, DC, Liu, T, MacLean, B, Mani, DR, Mansfield, E, Neubert, H, Paulovich, AG, Reiter, L, Vitek, O, Aebersold, R, Anderson, L, Bethem, R, Blonder, J, Boja, E, Botelho, J, Boyne, M, Bradshaw, RA, Burlingame, AL, Chan, D, Keshishian, H, Kuhn, E, Kinsinger, C, Lee, JSH, Lee, S-W, Moritz, R, Oses-Prieto, J, Rifai, N, Ritchie, J, Rodriguez, H, Srinivas, PR, R Townsend, R, Van Eyk, J, Whiteley, G, Wiita, A, Weintraub, S
JournalMol Cell Proteomics
Date Published2014 Mar
KeywordsAnimals, Biological Assay, Biology, Guidelines as Topic, Humans, Isotope Labeling, Mass Spectrometry, Medicine, Peptides, Proteomics, Reference Standards, Software

Adoption of targeted mass spectrometry (MS) approaches such as multiple reaction monitoring (MRM) to study biological and biomedical questions is well underway in the proteomics community. Successful application depends on the ability to generate reliable assays that uniquely and confidently identify target peptides in a sample. Unfortunately, there is a wide range of criteria being applied to say that an assay has been successfully developed. There is no consensus on what criteria are acceptable and little understanding of the impact of variable criteria on the quality of the results generated. Publications describing targeted MS assays for peptides frequently do not contain sufficient information for readers to establish confidence that the tests work as intended or to be able to apply the tests described in their own labs. Guidance must be developed so that targeted MS assays with established performance can be made widely distributed and applied by many labs worldwide. To begin to address the problems and their solutions, a workshop was held at the National Institutes of Health with representatives from the multiple communities developing and employing targeted MS assays. Participants discussed the analytical goals of their experiments and the experimental evidence needed to establish that the assays they develop work as intended and are achieving the required levels of performance. Using this "fit-for-purpose" approach, the group defined three tiers of assays distinguished by their performance and extent of analytical characterization. Computational and statistical tools useful for the analysis of targeted MS results were described. Participants also detailed the information that authors need to provide in their manuscripts to enable reviewers and readers to clearly understand what procedures were performed and to evaluate the reliability of the peptide or protein quantification measurements reported. This paper presents a summary of the meeting and recommendations.


Alternate JournalMol. Cell Proteomics
PubMed ID24443746
PubMed Central IDPMC3945918
Grant ListU24 CA160034 / CA / NCI NIH HHS / United States
R01HL096738 / HL / NHLBI NIH HHS / United States
P41 GM103481 / GM / NIGMS NIH HHS / United States
U24 CA159988 / CA / NCI NIH HHS / United States
R01 HL096738 / HL / NHLBI NIH HHS / United States
HHSN268201000033C / / PHS HHS / United States
P30 CA076292 / CA / NCI NIH HHS / United States
HHSN268201000033C / HL / NHLBI NIH HHS / United States
U24 CA160019 / CA / NCI NIH HHS / United States
U24 CA160035 / CA / NCI NIH HHS / United States
U24CA160034 / CA / NCI NIH HHS / United States