biostudies-literatureTsiklauri GBundesministerium für Bildung und ForschungElitenetzwerk Bayern4988-5000https://www.ebi.ac.uk/biostudies/studies/S-EPMC12501992biostudies-literatureUnknown24(10)Current applications of mass-spectrometry-based proteomics range from single-cell to body fluid analysis, each presenting very different demands regarding sensitivity or sample throughput. Additionally, the vast molecular complexity of proteomes and the massive dynamic range of protein concentrations in these biological systems require highly performant chromatographic separations in tandem with the high speed and sensitivity afforded by modern mass spectrometers. In this study, we focused on the chromatographic aspect and, more specifically, systematically evaluated proteome analysis performance across a wide range of chromatographic flow rates (0.3-50 μL/min) and associated column diameters using a Vanquish Neo HPLC coupled online to a Q Exactive HF-X mass spectrometer. Serial dilutions Journal of proteome researchHigh-Performance Proteomics Using Nano-, Capillary-, and Microflow Chromatographic Separations.PMC12501992F.6-M5613.6.K-NW-2021-411/1/103LW0243K16LW0243KProkofeva PTsiklauri GZheng RKabella NKuster BPynn CfalseHigh-Performance Proteomics Using Nano-, Capillary-, and Microflow Chromatographic Separations.Current applications of mass-spectrometry-based proteomics range from single-cell to body fluid analysis, each presenting very different demands regarding sensitivity or sample throughput. Additionally, the vast molecular complexity of proteomes and the massive dynamic range of protein concentrations in these biological systems require highly performant chromatographic separations in tandem with the high speed and sensitivity afforded by modern mass spectrometers. In this study, we focused on the chromatographic aspect and, more specifically, systematically evaluated proteome analysis performance across a wide range of chromatographic flow rates (0.3-50 μL/min) and associated column diameters using a Vanquish Neo HPLC coupled online to a Q Exactive HF-X mass spectrometer. Serial dilutions 2025-01-01T00:00:00Z2025 Oct2026-06-04T06:43:09.201Z2026-05-06T03:13:25.059ZS-EPMC125019924090372410.1021/acs.jproteome.5c00327