Proteomic analysis of complex protein mixtures using proteolytic digestion and liquid chromatography in combination with tandem mass spectrometry is a standard approach in biological studies. Data-dependent acquisition is used to automatically acquire tandem mass spectra of peptides eluting into the mass spectrometer. In more complicated mixtures, for example, whole cell lysates, data-dependent acquisition incompletely samples among the peptide ions present rather than acquiring tandem mass spectra for all ions available. We analyzed the sampling process and developed a statistical model to accurately predict the level of sampling expected for mixtures of a specific complexity. The model also predicts how many analyses are required for saturated sampling of a complex protein mixture. For a yeast-soluble cell lysate 10 analyses are required to reach a 95% saturation level on protein identifications based on our model. The statistical model also suggests a relationship between the level of sampling observed for a protein and the relative abundance of the protein in the mixture. We demonstrate a linear dynamic range over 2 orders of magnitude by using the number of spectra (spectral sampling) acquired for each protein.

• PMID: 15253663
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Comparative Study | Evaluation Study | Journal Article | Research Support, U.S. Gov't, Non-P.H.S. | Research Support, U.S. Gov't, P.H.S.

Authors

Liu H, Sadygov RG, Yates JR

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