PXD007894

PXD007894 is an original dataset announced via ProteomeXchange.

Title	Absolute quantitation of proteins by two-point quantitation using dimethylated proteotypic peptides
Description	Absolute targeted proteomics typically employs known amounts of synthetic stable isotopically labeled peptides which are mixed with the analyte and analysed by LC-MS. In order to obtain accurate data, we evaluated the use of two different stable isotopes of the same peptide as spike-in for absolute quantification. For this purpose, peptide labeling by reductive amination was applied, which is a mild reaction for dimethylation of amine groups with very high yield and very little side reactions. Furthermore, three different forms can be generated with e.g., light and heavy labels for spike-in peptides, and medium label for endogenous peptides. The addition of the dimethylated proteotypic peptide in two different versions allows to determine the accuracy of the quantification of endogenous peptides using the relative ratio of the intensities of the two internal standards. The method was studied with peptides of apolipoprotein A-I, apolipoprotein B-100, and leucine-rich alpha-2-glycoprotein without and with serum. In serum, the endogenous protein concentrations were accurately measured across four orders of magnitude by the two-point quantitation method. Less than 20% of coefficient of variation (CV) values and strong correlation with R2 of 0.99 across three analytical replicates was observed. The results showed that the accuracy to determine the amount of endogenous peptides can be validated using two references against each other, which cannot be obtained by one-point quantitation. Because of the significant lower costs than synthetic stable isotopically labeled peptides, this approach might be particularly interesting for the absolute quantitation of multiple proteins.
HostingRepository	PRIDE
AnnounceDate	2018-11-27
AnnouncementXML	Submission_2018-11-27_04:27:37.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Trung Tran
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	No PTMs are included in the dataset
Instrument	Q Exactive

Revision	Datetime	Status	ChangeLog Entry
0	2017-10-03 01:33:18	ID requested
⏵ 1	2018-11-27 04:27:38	announced

Tran TT, Bollineni RC, Koehler CJ, Thiede B, Absolute two-point quantification of proteins using dimethylated proteotypic peptides. Analyst, 143(18):4359-4365(2018) [pubmed]

submitter keyword: absolute quantitation, dimethylation

mass spectrometry, parallel reaction monitoring, targeted proteomics

Bernd Thiede
contact affiliation	Department of Biosciences, University of Oslo, P.O. Box 1066 Blindern, 0316 Oslo, Norway, Tel.: +47-22840533
contact email	bernd.thiede@ibv.uio.no
lab head
Trung Tran
contact affiliation	University of Oslo
contact email	trung.tran@ibv.uio.no
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/2018/11/PXD007894

PRIDE project URI

• PRIDE
  1. PXD007894
     1. Label: PRIDE project
     2. Name: Absolute quantitation of proteins by two-point quantitation using dimethylated proteotypic peptides