Ultra-fast label-free quantification and comprehensive proteome coverage with narrow-window data-independent acquisition
Ulises H. Guzmán, Ana Martínez‐Val, Zilu Ye, Eugen Damoc, Tabiwang N. Arrey, Anna Pashkova, Santosh Renuse, Eduard Denisov, J. Petzoldt, Amelia C. Peterson, Florian Harking, Ole Østergaard, Rasmus Rydbirk, Susana Aznar, Hamish Stewart, Yue Xuan, Daniel Hermanson, Stevan Horning, Christian Hock, Alexander Makarov, Vlad 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.