PXD058591

PXD058591 is an original dataset announced via ProteomeXchange.

Title	SPE-CZE-MS Quantifies Zeptomole Amounts of Phosphorylated Peptides
Description	Capillary zone electrophoresis (CZE) is gaining attention in the field of single-cell proteomics for its ultra-low-flow and high-resolution separation abilities. Even more sample-limited yet rich in biological information are phosphoproteomics experiments, as the phosphoproteome composes only a fraction of the whole cellular proteome and preparation for this kind of analysis requires an additional enrichment step. Rapid analysis, high sensitivity, and maximization of sample utilization is paramount for single-cell analysis. Some challenges of coupling CZE analysis with mass spectrometry analysis (MS) of complex mixtures include 1. sensitivity due to volume loading limitations of CZE and 2. incompatibility of MS duty cycles with electropherographic timescales, especially for rapid (<20 min) analyses. Here, we address these two challenges as applied to single-cell equivalent phosphoproteomics experiments by interfacing a microchip-based CZE device integrated with a solid-phase-extraction (SPE) bed with the Orbitrap Astral mass spectrometer. The SPE bed theoretically circumvents the volume limitations that thwart detection sensitivity of classical CZE by allowing analyte capture and concentration before electrokinetic focusing and separation. Further, we leverage the high sampling rates of the Orbitrap Astral mass spectrometer to take full advantage of the fast and high-resolution SPE-CZE separations which historically pose challenges for typical MS scan speeds. Using 225 phosphorylated peptide standards and phosphorylated peptide-enriched mouse brain tissue, we investigate microchip-based SPE-CZE functionality, quantitative performance, and complementarity to nano-LC-MS analysis. We characterize important device characteristics such as volume and mass loading tolerances as well as compatibility with state-of-the-art mass spectrometry methods. Finally, we highlight unique SPE-CZE separation mechanisms that can empower fit-for-purpose applications in single-cell-equivalent phosphoproteomics.
HostingRepository	MassIVE
AnnounceDate	2025-05-08
AnnouncementXML	Submission_2025-05-08_02:51:13.308.xml
DigitalObjectIdentifier
ReviewLevel	Non peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Lia R Serrano
SpeciesList	scientific name: Mus musculus; common name: house mouse; NCBI TaxID: 10090;
ModificationList	Phospho
Instrument	Orbitrap Astral

Revision	Datetime	Status	ChangeLog Entry
0	2024-12-04 18:14:38	ID requested
⏵ 1	2025-05-08 02:51:13	announced

no publication

submitter keyword: Phosphoproteomics, SPE-CZE, DatasetType:Proteomics

Joshua Coon
contact affiliation	University of Wisconsin - Madison
contact email	jcoon@chem.wisc.edu
lab head
Lia R Serrano
contact affiliation	University of Wisconsin Madison
contact email	lrserrano@wisc.edu
dataset submitter

MassIVE dataset URI

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