Abstract
Quantitative proteomics employing mass spectrometry is an indispensable tool in life science research. Targeted proteomics has emerged as a powerful approach for reproducible quantification but is limited in the number of proteins quantified. SWATH-mass spectrometry consists of data-independent acquisition and a targeted data analysis strategy that aims to maintain the favorable quantitative characteristics (accuracy, sensitivity, and selectivity) of targeted proteomics at large scale. While previous SWATH-mass spectrometry studies have shown high intra-lab reproducibility, this has not been evaluated between labs. In this multi-laboratory evaluation study including 11 sites worldwide, we demonstrate that using SWATH-mass spectrometry data acquisition we can consistently detect and reproducibly quantify >4000 proteins from HEK293 cells. Using synthetic peptide dilution series, we show that the sensitivity, dynamic range and reproducibility established with SWATH-mass spectrometry are uniformly achieved. This study demonstrates that the acquisition of reproducible quantitative proteomics data by multiple labs is achievable, and broadly serves to increase confidence in SWATH-mass spectrometry data acquisition as a reproducible method for large-scale protein quantification.
Highlights
Quantitative proteomics employing mass spectrometry is an indispensable tool in life science research
A physiologically relevant fold change step, and to cover a large linear dynamic range in a relatively small number of samples that could be analyzed in a 24 h period, we elected to partition the stable isotopelabeled standard (SIS) peptides into five groups (A–E), each containing six peptides
The importance of quantitative proteomics in clinical and basic research is expanding rapidly because proteins provide a direct insight into the biochemical state of the cell
Summary
Quantitative proteomics employing mass spectrometry is an indispensable tool in life science research. The question of whether specific data acquisition methods and platforms are capable of generating reproducible results is best addressed by inter-laboratory studies, where samples of known composition and quality are analyzed across different settings. Such studies have been reported for various “omics” technologies, including RNA-seq and microarray techniques, with varying results[4, 5]. The number of proteins measured is restricted (usually to ~100 per injection), limiting its utility for many applications
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
