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Updated: Jul 4, 2025
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Deep Proteome Profiling by Isobaric Labeling, Extensive Liquid Chromatography, Mass Spectrometry, and Software-assisted Quantification
Published on: November 15, 2017
12.0K
Ultra-fast label-free quantification and comprehensive proteome coverage with narrow-window data-independent
Ulises H Guzman1, Ana Martinez-Val1, Zilu Ye1,2
1Novo Nordisk Foundation Center for Protein Research, University of Copenhagen, Copenhagen, Denmark.
Nature Biotechnology
|February 1, 2024
Summary
A new narrow-window data-independent acquisition (nDIA) strategy enhances proteome characterization speed and depth. This advanced mass spectrometry approach achieves higher protein coverage for faster, more comprehensive proteomic analysis.
Area of Science:
- Proteomics
- Mass Spectrometry
- Analytical Chemistry
Background:
- Mass spectrometry (MS)-based proteomics is crucial for characterizing proteomes.
- Current methods face limitations in speed and reproducibility.
- Bridging data-dependent and data-independent acquisition is an ongoing challenge.
Purpose of the Study:
- To introduce and validate the narrow-window data-independent acquisition (nDIA) strategy.
- To enhance the speed and depth of proteome profiling.
- To demonstrate the capabilities of a novel quadrupole Orbitrap mass spectrometer with an asymmetric track lossless (Astral) analyzer.
Main Methods:
- Utilized narrow-window (2-Th) isolation windows with high-resolution MS1 scans.
- Employed parallel tandem MS (MS/MS) scans at >200 Hz using the Astral analyzer.
- Integrated high resolving power, sensitivity, and low-ppm mass accuracy.
Main Results:
- Achieved profiling of >100 yeast proteomes or 48 human proteomes per day.
- Enabled deep proteome coverage (~10,000 protein groups in 30 min; ~7,000 proteins in 5 min).
- Demonstrated 3x higher protein coverage compared to state-of-the-art MS methods.
- Quantified over 14,000 protein groups in a single 30-min run with high precision and accuracy.
Conclusions:
- The nDIA strategy significantly advances proteomic analysis speed and coverage.
- The Astral analyzer is key to achieving high-throughput, high-depth proteome profiling.
- nDIA offers a powerful approach for comprehensive and quantitative proteome characterization.
