PXD072114

PXD072114 is an original dataset announced via ProteomeXchange.

Title	Limited impact of column chemistry and length on proteome coverage under high-speed DIA
Description	The evolution of mass spectrometry-based proteomics has been driven by continuous advances in instrumentation, sample preparation, and data acquisition strategies. While chromatographic separation has historically been considered a critical bottleneck in achieving comprehensive proteome coverage, recent developments in ultra-fast mass spectrometry acquisition fundamentally challenge this paradigm. We investigated whether traditional chromatographic optimization principles established during the early era of proteomics remain essential in contemporary workflows. Using five distinct stationary phases, C18 chemistries, C8, and Phenyl-Hexyl, across eight column lengths (40-140 mm), we evaluated proteome identification performance using state-of-the-art data-independent acquisition on the Orbitrap Astral mass spectrometer with HeLa tryptic digests. Despite substantial chromatographic differences in selectivity and peak characteristics that would have profoundly influenced analytical outcomes in earlier instrumentation generations, we observed remarkably convergent proteome coverage metrics. All C18 and C8 phases consistently achieved over 150,000 precursor and approximately 9,000 protein group identifications, regardless of column length variations. While distinct selectivity fingerprints persisted across chemistries, these chromatographic differences did not translate into meaningful variations in bulk identification depth under high-speed acquisition conditions exceeding 200 Hz. From these findings we conclude that the analytical bottleneck has fundamentally shifted from chromatographic resolution to mass spectrometric sampling efficiency, where comprehensive peptide identification is now achieved through advanced spectral deconvolution rather than physical separation alone. This paradigmatic shift suggests that method development priorities in modern proteomics should evolve beyond traditional separation optimization to emphasize operational robustness, analytical throughput, and systematic reproducibility for routine applications.
HostingRepository	PRIDE
AnnounceDate	2026-06-23
AnnouncementXML	Submission_2026-06-22_20:14:58.632.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Mario Oroshi
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: NEWT:9606; scientific name: Mus musculus (Mouse); NCBI TaxID: NEWT:10090; scientific name: Arabidopsis thaliana (Mouse-ear cress); NCBI TaxID: NEWT:3702;
ModificationList	iodoacetamide derivatized residue
Instrument	Orbitrap Astral

Revision	Datetime	Status	ChangeLog Entry
0	2025-12-17 13:45:16	ID requested
⏵ 1	2026-06-22 20:14:59	announced

Dataset with its publication pending

submitter keyword: Column performance, Chromatographic selectivity, Method development,LC-MS proteomics, Orbitrap Astral

Dr. Johannes Bruno Müller-Reif
contact affiliation	Project Group Leader Dept. of Proteomics and Signal Transduction Max Planck Institute of Biochemistry
contact email	jomueller@biochem.mpg.de
lab head
Mario Oroshi
contact affiliation	Proteomics
contact email	oroshi@biochem.mpg.de
dataset submitter

Dataset FTP location NOTE: Most web browsers have now discontinued native support for FTP access within the browser window. But you can usually install another FTP app (we recommend FileZilla) and configure your browser to launch the external application when you click on this FTP link. Or otherwise, launch an app that supports FTP (like FileZilla) and use this address: ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD072114

PRIDE project URI

• PRIDE
  1. PXD072114
     1. Label: PRIDE project
     2. Name: Limited impact of column chemistry and length on proteome coverage under high-speed DIA