Journal
ANALYTICAL CHEMISTRY
Volume 92, Issue 6, Pages 4326-4333Publisher
AMER CHEMICAL SOC
DOI: 10.1021/acs.analchem.9b05095
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Funding
- Russian Science Foundation [19-74-00123]
- European Research Council (ERC) under the European Union [646603]
- VILLUM Center for Bioanalytical Sciences at the University of Southern Denmark
- European Research Council (ERC) [646603] Funding Source: European Research Council (ERC)
- Russian Science Foundation [19-74-00123] Funding Source: Russian Science Foundation
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Proteome characterization relies heavily on tandem mass spectrometry (MS/MS) and is thus associated with instrumentation complexity, lengthy analysis time, and limited duty cycle. It was always tempting to implement approaches that do not require MS/MS, yet they were constantly failing to achieve a meaningful depth of quantitative proteome coverage within short experimental times, which is particularly important for clinical or biomarker-discovery applications. Here, we report on the first successful attempt to develop a truly MS/MS-free method, DirectMS1, for bottom-up proteomics. The method is compared with the standard MS/MS-based data-dependent acquisition approach for proteome-wide analysis using 5 min LC gradients. Specifically, we demonstrate identification of 1 000 protein groups for a standard HeLa cell line digest. The amount of loaded sample was varied in a range from 1 to 500 ng, and the method demonstrated 10-fold higher sensitivity. Combined with the recently introduced Diffacto approach for relative protein quantification, DirectMS1 outperforms most popular MS/MS-based label-free quantitation approaches because of significantly higher protein sequence coverage.
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