PXD000705

PXD000705 is an original dataset announced via ProteomeXchange.

Title	Extending proteome coverage through an on-line 2D SCX-RP ultra-high pressure system
Description	Ultra high pressure liquid chromatography (UHPLC) systems combined with state of the art mass spectrometers have pushed the limit of deep proteome sequencing to new heights making it possible to identify thousands of proteins in a single LC/MS experiment within a few hours. However, the relationship between gradient length and the number of proteins identified is not linear and as gradient time increases to several hours, performance gain diminishes. Proteome coverage can be extended using 2-dimensional chromatography, but traditionally this comes at the expense of sample losses and much longer analysis times. Here, we asked the question whether a fast and sensitive online 2D SCX-RP UHPLC-MS/MS workflow, could compete with the current 1D long gradient analysis, making total analysis time- versus proteome coverage and sample used as benchmark parameters. Our new automated 2D-LC/MS system is robust and easy to use, consisting of a homemade SCX column, a trap column and a 50 cm analytical EASY-Spray column. We benchmark the system using small amounts of a human cell lysate digest. The 2D SCX-RP UHPLC-MS/MS workflow allowed us to identify 56600 unique peptides and 5958 proteins in a total analysis time of just over 8 hours. On the same system a 1D RP UHPLC-MS/MS workflow gave only 22000 peptides and 4200 unique proteins in 6 hours analysis time, increases of 157% and 42% at the peptide and protein level, respectively. These and other data reported here reveal that with this fast online SCX-RP UHPLC-MS/MS workflow proteome coverage can be substantially extended without compromising too much analysis time and sample usage
HostingRepository	PRIDE
AnnounceDate	2014-10-01
AnnouncementXML	Submission_2016-06-01_01:24:22.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Fabio Marino
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	monohydroxylated residue; acetylated residue; iodoacetamide derivatized residue; phosphorylated residue
Instrument	Q Exactive

Revision	Datetime	Status	ChangeLog Entry
0	2014-01-22 06:45:11	ID requested
1	2014-09-30 04:00:29	announced
2	2014-10-01 00:30:47	announced	Updated project metadata.
⏵ 3	2016-06-01 01:24:24	announced	Updated project metadata.

Marino F, Cristobal A, Binai NA, Bache N, Heck AJ, Mohammed S, Characterization and usage of the EASY-spray technology as part of an online 2D SCX-RP ultra-high pressure system. Analyst, 139(24):6520-8(2014) [pubmed]

curator keyword: Technical

submitter keyword: 2D LC-MS/MS, Proteomics

Albert J.R. Heck
contact affiliation	Biomolecular Mass Spectrometry and Proteomics groups, Utrecht University
contact email	a.j.r.heck@uu.nl
lab head
Fabio Marino
contact affiliation	Utrecht University
contact email	F.Marino@uu.nl
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/2014/09/PXD000705

PRIDE project URI

• PRIDE
  1. PXD000705
     1. Label: PRIDE project
     2. Name: Extending proteome coverage through an on-line 2D SCX-RP ultra-high pressure system