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PXD031277-1

PXD031277 is an original dataset announced via ProteomeXchange.

Dataset Summary
TitleOptimal analytical strategies for sensitive and quantitative phosphoproteomics using TMT-based multiplexing
DescriptionIn large-scale quantitative mass spectrometry (MS)-based phosphoproteomics, isobaric labeling with tandem mass tags (TMTs) coupled with offline high-pH reversed-phase peptide chromatographic fractionation maximizes depth of coverage. To investigate to what extent limited sample amounts affects sensitivity and dynamic range of the analysis due to sample losses, we benchmarked TMT-based peptide fractionation strategies against single-shot (SS) label-free approach with data independent acquisition (DIA), for different peptide input per sample. To systematically examine how peptide input amounts influence TMT-fractionation approaches in a phosphoproteomics workflow, we compared two different high-pH reverse-phase fractionation strategies, microflow (MF) and stage-tip fractionation (STF), while scaling the peptide input amount down from 12.5 μg to 1 μg per sample. Our results indicate that, for input amounts higher than 5 μg per sample, TMT labeling, followed by microflow fractionation and phospho-enrichment (MF), achieves the deepest phosphoproteome coverage, even compared to SS DIA analysis. Conversely, stage-tip fractionation of enriched phosphopeptides (STF) is optimal for lower amounts, below 5 μg/peptide per sample. As a result, we provide a decision tree to help phosphoproteomics users to choose the best workflow as a function of on sample amount.
HostingRepositoryPRIDE
AnnounceDate2022-06-28
AnnouncementXMLSubmission_2022-06-28_10:30:13.389.xml
DigitalObjectIdentifier
ReviewLevelPeer-reviewed dataset
DatasetOriginOriginal dataset
RepositorySupportUnsupported dataset by repository
PrimarySubmitterClaire Koenig
SpeciesList scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationListphosphorylated residue
InstrumentOrbitrap Exploris 480
Dataset History
RevisionDatetimeStatusChangeLog Entry
02022-01-26 12:35:16ID requested
12022-06-28 10:30:13announced
22023-11-14 08:54:50announced2023-11-14: Updated project metadata.
Publication List
10.1002/PMIC.202100245;
Keyword List
submitter keyword: Phosphoproteomics, Isobaric labeling, Tandem mass tags, high-pH fractionation, optimization
Contact List
Jesper V. Olsen
contact affiliationGroup leader, Professor University of Copenhagen Faculty of Health and Medical Sciences Blegdamsvej 3B København N
contact emailjesper.olsen@cpr.ku.dk
lab head
Claire Koenig
contact affiliationNovo Nordisk Foundation Center for Protein Research, University of Copenhagen
contact emailclaire.koenig@cpr.ku.dk
dataset submitter
Full Dataset Link List
Dataset FTP location
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