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Options and considerations when selecting a quantitative proteomics strategy

Abstract

The vast majority of proteomic studies to date have relied on mass spectrometric techniques to identify, and in some cases quantify, peptides that have been generated by proteolysis. Current approaches differ in the types of instrument used, their performance profiles, the manner in which they interface with biological research strategies, and their reliance on and use of prior information. Here, we consider the three main mass spectrometry (MS)-based proteomic approaches used today: shotgun (or discovery), directed and targeted strategies. We discuss the principles of each technique, their strengths and weaknesses and the dependence of their performance profiles on the composition of the biological sample. Our goal is to provide a rational framework for selecting strategies optimally suited to address the specific research issue under consideration.

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Figure 1: Workflow of a typical proteomic experiment.
Figure 2: Workflow of a discovery proteomic experiment.
Figure 3: Workflow of a directed proteomic experiment.
Figure 4: Workflow of a targeted proteomic experiment.
Figure 5: A representation of the desired characteristics of a proteomic experiment.
Figure 6: Performance profiles of the shotgun or discovery (a), directed (b) and targeted (c) proteomic methods.
Figure 7: Effect on biochemical background on quantification by the shotgun (discovery), directed and targeted proteomics strategies.
Figure 8: Effect of background on the detection of a peptide mixture in various concentrations (arbitrary units).
Figure 9: Isotopically labeled internal standards can be added at various stages in quantitative proteomics experiments.

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Acknowledgements

We thank P. Picotti, A. Schmidt and N. Selevsek for the preparation of figures. The Swiss National Science Foundation (grant no. 3100AO-107679), SystemsX.ch, the Swiss initiative for systems biology and the European Research Council are acknowledged for financial support.

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Correspondence to Bruno Domon or Ruedi Aebersold.

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Domon, B., Aebersold, R. Options and considerations when selecting a quantitative proteomics strategy. Nat Biotechnol 28, 710–721 (2010). https://doi.org/10.1038/nbt.1661

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