Ultra-fast label-free quantification and comprehensive proteome coverage with narrow-window data-independent acquisition

Ulises H Guzman, Ana Martinez-Val, Zilu Ye, Eugen Damoc, Tabiwang N Arrey, Anna Pashkova, Santosh Renuse, Eduard Denisov, Johannes Petzoldt, Amelia C Peterson, Florian Harking, Ole Østergaard, Rasmus Rydbirk, Susana Aznar, Hamish Stewart, Yue Xuan, Daniel Hermanson, Stevan Horning, Christian Hock, Alexander MakarovVlad Zabrouskov, Jesper V Olsen

Abstract

Mass spectrometry (MS)-based proteomics aims to characterize comprehensive proteomes in a fast and reproducible manner. Here we present the narrow-window data-independent acquisition (nDIA) strategy consisting of high-resolution MS1 scans with parallel tandem MS (MS/MS) scans of ~200 Hz using 2-Th isolation windows, dissolving the differences between data-dependent and -independent methods. This is achieved by pairing a quadrupole Orbitrap mass spectrometer with the asymmetric track lossless (Astral) analyzer which provides >200-Hz MS/MS scanning speed, high resolving power and sensitivity, and low-ppm mass accuracy. The nDIA strategy enables profiling of >100 full yeast proteomes per day, or 48 human proteomes per day at the depth of ~10,000 human protein groups in half-an-hour or ~7,000 proteins in 5 min, representing 3× higher coverage compared with current state-of-the-art MS. Multi-shot acquisition of offline fractionated samples provides comprehensive coverage of human proteomes in ~3 h. High quantitative precision and accuracy are demonstrated in a three-species proteome mixture, quantifying 14,000+ protein groups in a single half-an-hour run.

Original languageEnglish
JournalNature Biotechnology
ISSN1087-0156
DOIs
Publication statusE-pub ahead of print - 1 Feb 2024

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