期刊
ANALYTICAL CHEMISTRY
卷 94, 期 4, 页码 2016-2022出版社
AMER CHEMICAL SOC
DOI: 10.1021/acs.analchem.1c03782
关键词
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资金
- Ministry of Science and Higher Education of the Russian Federation (Skolkovo Institute of Science and Technology) [075-10-2019-083]
- Genome Canada [GTP: 264PRO]
- Segal McGill Chair in Molecular Oncology at McGill University (Montreal, Quebec, Canada)
- Warren Y. Soper Charitable Trust
- Alvin Segal Family Foundation
Mass spectrometry-based quantitative proteomic methods, such as prm-PASEF, are important tools for protein biomarker discovery and validation. This study demonstrated the use of prm-PASEF for absolute protein quantitation in human plasma, with improved sensitivity and a broad dynamic range. The prm-PASEF method showed excellent correlation with MRM, highlighting its suitability for absolute quantitative proteomics.
Mass spectrometry (MS)-based quantitative proteomic methods have become some of the major tools for protein biomarker discovery and validation. The recently developed parallel reaction monitoring-parallel accumulation-serial fragmentation (prm-PASEF) approach on a Bruker timsTOF Pro mass spectrometer allows the addition of ion mobility as a new dimension to LC-MS-based proteomics and increases proteome coverage at a reduced analysis time. In this study, a prm-PASEF approach was used for the multiplexed absolute quantitation of proteins in human plasma using isotope-labeled peptide standards for 125 plasma proteins, over a broad (10(4)-10(6)) dynamic range. Optimization of LC and MS parameters, such as accumulation time and collision energy, resulted in improved sensitivity for more than half of the targets (73 out of 125 peptides) by increasing the signal-tonoise ratio by a factor of up to 10. Overall, 41 peptides showed up to a 2-fold increase in sensitivity, 25 peptides showed up to a 5-fold increase in sensitivity, and 7 peptides showed up to a 10-fold increase in sensitivity. Implementation of the prm-PASEF method allowed absolute protein quantitation (down to 1.13 fmol) in human plasma samples. A comparison of the concentration values of plasma proteins determined by MRM on a QTRAP instrument and by prm-PASEF on a timsTOF Pro revealed an excellent correlation (R-2 = 0.97) with a slope of close to 1 (0.99), demonstrating that prm-PASEF is well suited for absolute quantitative proteomics.
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