PXD025387

PXD025387 is an original dataset announced via ProteomeXchange.

Title	An automated workflow for label-free and multiplexed single cell proteomics sample preparation at unprecedented sensitivity.
Description	The analysis of single cell proteomes has recently become a viable complement to transcript and genomics studies. Proteins are the main driver of cellular functionality and mRNA levels are often an unreliable proxy of such. Therefore, the global analysis of the proteome is essential to study cellular identities. Both multiplexed and label-free mass spectrometry-based approaches with single cell resolution have lately attributed surprising heterogeneity to believed homogenous cell populations. Even though specialized experimental designs and instrumentation have demonstrated remarkable advances, the efficient sample preparation of single cells still lacks behind. Here, we introduce the proteoCHIP, a universal option for single cell proteomics sample preparation at surprising sensitivity and throughput. The automated processing using a commercial system combining single cell isolation and picoliter dispensing, the cellenONE®, allows to reduce final sample volumes to low nanoliters submerged in a hexadecane layer simultaneously eliminating error prone manual sample handling and overcoming evaporation. With this specialized workflow we achieved around 1,000 protein groups per analytical run at remarkable reporter ion signal to noise while reducing or eliminating the carrier proteome. We identified close to 2,000 protein groups across 158 multiplexed single cells from two highly similar human cell types and clustered them based on their proteome. In-depth investigation of regulated proteins readily identified one of the main drivers for tumorigenicity in this cell type. Our workflow is compatible with all labeling reagents, can be easily adapted to custom workflows and is a viable option for label-free sample preparation. The specialized proteoCHIP design allows for the direct injection of label-free single cells via a standard autosampler resulting in the recovery of 30% more protein groups compared to samples transferred to PEG coated vials. We therefore are confident that our versatile, sensitive, and automated sample preparation workflow will be easily adoptable by non-specialized groups and will drive biological applications of single cell proteomics.
HostingRepository	PRIDE
AnnounceDate	2021-04-19
AnnouncementXML	Submission_2021-04-19_02:11:42.582.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Claudia Ctortecka
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	TMT6plex-126 reporter+balance reagent acylated residue; acetylated residue; monohydroxylated residue
Instrument	Orbitrap Exploris 480

Revision	Datetime	Status	ChangeLog Entry
0	2021-04-14 01:57:21	ID requested
⏵ 1	2021-04-19 02:11:42	announced

Dataset with its publication pending

submitter keyword: single cell proteomics, proteoCHIP, multiplexed, label-free

Karl Mechtler
contact affiliation	Research Institute of Molecular Pathology (IMP), Vienna BioCenter (VBC), Campus-Vienna-Biocenter 1, 1030 Vienna, Austria.
contact email	karl.mechtler@imp.ac.at
lab head
Claudia Ctortecka
contact affiliation	IMP
contact email	claudia.ctortecka@imp.ac.at
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/2021/04/PXD025387

PRIDE project URI

• PRIDE
  1. PXD025387
     1. Label: PRIDE project
     2. Name: An automated workflow for label-free and multiplexed single cell proteomics sample preparation at unprecedented sensitivity.