PXD005945

PXD005945 is an original dataset announced via ProteomeXchange.

Title	Deep LC-MS/MS analysis of the proteome from six cell lines
Description	Mass spectrometry is a rational orthogonal method for antibody-based assays, but implementation of MS in the validation pipeline of antibody manufacturers is hampered by the high cost and low throughput. Here we present a rapid method for antibody validation based on denaturing gel electrophoresis of biotinylated cell lysates (PAGE) followed by mass spectrometry (MS) and antibody array analysis (MAP). The first step, PAGE, produces 12 fractions containing proteins of increasing molecular weight. The fractions are analyzed in parallel by MS and MAP. Antibodies to be tested are immobilized on color coded polymer beads to create antibody arrays, which can comprise up to several thousand various antibodies. MS data provide definite protein identifications in each fraction, creating a reference for antibody reactivity patterns obtained via MAP. The method employs automated software to compare both datasets and provide validation data for each antibody tested. Due to the high-throughput nature of the assay we were able to screen several thousands of antibodies against six different cell lines. The differences in protein expression between the cell lines provide an additional control of antibody specificity. Using PAGE-MAP it is possible to screen and validate thousands of antibodies in a matter of weeks. Moreover, antibodies are tested under standardized conditions, which allows for direct comparison of their performance.
HostingRepository	PRIDE
AnnounceDate	2024-10-22
AnnouncementXML	Submission_2024-10-22_04:36:15.178.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Krzysztof Sikorski
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	isotope labeled residue
Instrument	Q Exactive

Revision	Datetime	Status	ChangeLog Entry
0	2017-02-21 05:46:33	ID requested
1	2018-07-19 04:48:49	announced
⏵ 2	2024-10-22 04:36:15	announced	2024-10-22: Updated project metadata.

Dataset with its publication pending

curator keyword: Biological

submitter keyword: PAGE-MAP, GelFree,on-bead digestion, antibody validation, LC-MS/MS

Fridtjof Lund-Johansen
contact affiliation	Dept. of Immunology, Oslo University Hospital Rikshospitalet, Oslo, Norway
contact email	fridtjol@gmail.com
lab head
Krzysztof Sikorski
contact affiliation	K.G. Jebsen Center for Cancer Immunotherapy, Oslo, Norway
contact email	kristof.sikorski@gmail.com
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/2018/07/PXD005945

PRIDE project URI

• PRIDE
  1. PXD005945
     1. Label: PRIDE project
     2. Name: Deep LC-MS/MS analysis of the proteome from six cell lines