PXD060786

PXD060786 is an original dataset announced via ProteomeXchange.

Title	Development of an Adaptive, Economical, and Easy-to-Use SP3-TMT Automated Sample Preparation Workflow for Quantitative Proteomics
Description	Liquid handling robots have been developed to automate various steps of the bottom-up proteomics workflow, however, protocols for the generation of isobarically labeled peptides remain limited. Existing methods often require costly specialty devices and are constrained by fixed workflows. To address this, we developed a cost-effective, flexible, automated sample preparation protocol for TMT-labeled peptides using the Biomek i5 liquid handler. Our approach leverages Single-Pot Solid-Phase-Enhanced Sample Preparation (SP3) with paramagnetic beads to streamline protein cleanup and digestion. The protocol also allows for adjustment of trypsin concentration and peptide-to-TMT ratio to increase throughput and reduce costs, respectively. We compared our automated and manual 18-plex TMT-Pro labeling workflows by monitoring select protein markers of the Unfolded Protein Response (UPR) in pharmacologically activatable, engineered cell lines. Overall, the automated protocol demonstrated equivalent performance in peptide and protein identifications, digestion and labeling efficiency, and an enhancement in the dynamic range of TMT quantifications. Compared to the manual method, the Biomek protocol significantly reduces hands-on time and minimizes sample handling errors. The 96-well format additionally allows for the number of TMT reactions to be scaled up quickly without a significant increase in user interaction. Our optimized automated workflow enhances throughput, reproducibility, and cost-effectiveness, making it a valuable tool for high-throughput proteomics studies.
HostingRepository	PRIDE
AnnounceDate	2025-06-16
AnnouncementXML	Submission_2025-06-15_16:18:59.693.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Jake Hermanson
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	No PTMs are included in the dataset
Instrument	Orbitrap Exploris 480

Revision	Datetime	Status	ChangeLog Entry
0	2025-02-13 13:24:17	ID requested
⏵ 1	2025-06-15 16:19:00	announced

10.1021/acs.jproteome.5c00124;

Hermanson JN, Barny LA, Plate L, Development of an Adaptive, Economical, and Easy-to-Use SP3-TMT Automated Sample Preparation Workflow for Quantitative Proteomics. J Proteome Res, 24(6):2996-3006(2025) [pubmed]

submitter keyword: Automation, Sample Preperation, Unfolded Protein Response and Tandem Mass Tags

Lars Plate
contact affiliation	Department of Biological Sciences, Department of Chemistry, Vanderbilt University. Department of Pathology, Microbiology and Immunology, Vandberbilt University Medical Center
contact email	lars.plate@vanderbilt.edu
lab head
Jake Hermanson
contact affiliation	Vanderbilt University
contact email	jake.hermanson@vanderbilt.edu
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/2025/06/PXD060786

PRIDE project URI

• PRIDE
  1. PXD060786
     1. Label: PRIDE project
     2. Name: Development of an Adaptive, Economical, and Easy-to-Use SP3-TMT Automated Sample Preparation Workflow for Quantitative Proteomics