Click here to close
Hello! We notice that you are using Internet Explorer, which is not supported by Xenbase and may cause the site to display incorrectly.
We suggest using a current version of Chrome,
FireFox, or Safari.
???displayArticle.abstract???
Isobaric labeling strategies, such as isobaric tags for relative and absolute quantitation (iTRAQ) or tandem mass tags (TMT), have promised to dramatically increase the power of quantitative proteomics. However, when applied to complex mixtures, both the accuracy and precision are undermined by interfering peptide ions that coisolate and cofragment with the target peptide. Additional gas-phase isolation steps, such as proton-transfer ion-ion reactions (PTR) or higher-order MS3 scans, can almost completely eliminate this problem. Unfortunately, these methods come at the expense of decreased acquisition speed and sensitivity. Here we present a method that allows accurate quantification of TMT-labeled peptides at the MS2 level without additional ion purification. Quantification is based on the fragment ion cluster that carries most of the TMT mass balance. In contrast to the use of low m/z reporter ions, the localization of these complement TMT (TMT(C)) ions in the spectrum is precursor-specific; coeluting peptides do not generally affect the measurement of the TMT(C) ion cluster of interest. Unlike the PTR or MS3 strategies, this method can be implemented on a wide range of high-resolution mass spectrometers like the quadrupole Orbitrap instruments (QExactive). A current limitation of the method is that the efficiency of TMT(C) ion formation is affected by both peptide sequence and peptide ion charge state; we discuss potential routes to overcome this problem. Finally, we show that the complement reporter ion approach allows parallelization of multiplexed quantification and therefore holds the potential to multiply the number of distinct peptides that can be quantified in a given time frame.
Beausoleil,
Large-scale characterization of HeLa cell nuclear phosphoproteins.
2004, Pubmed
Beausoleil,
Large-scale characterization of HeLa cell nuclear phosphoproteins.
2004,
Pubmed
Beck,
The quantitative proteome of a human cell line.
2011,
Pubmed
Conrads,
Quantitative analysis of bacterial and mammalian proteomes using a combination of cysteine affinity tags and 15N-metabolic labeling.
2001,
Pubmed
Deeb,
Super-SILAC allows classification of diffuse large B-cell lymphoma subtypes by their protein expression profiles.
2012,
Pubmed
Dephoure,
Hyperplexing: a method for higher-order multiplexed quantitative proteomics provides a map of the dynamic response to rapamycin in yeast.
2012,
Pubmed
Gillet,
Targeted data extraction of the MS/MS spectra generated by data-independent acquisition: a new concept for consistent and accurate proteome analysis.
2012,
Pubmed
Gygi,
Quantitative analysis of complex protein mixtures using isotope-coded affinity tags.
1999,
Pubmed
Huttlin,
A tissue-specific atlas of mouse protein phosphorylation and expression.
2010,
Pubmed
Matthiesen,
Methods and algorithms for relative quantitative proteomics by mass spectrometry.
2010,
Pubmed
Mertins,
iTRAQ labeling is superior to mTRAQ for quantitative global proteomics and phosphoproteomics.
2012,
Pubmed
Nagaraj,
Deep proteome and transcriptome mapping of a human cancer cell line.
2011,
Pubmed
Olsen,
Quantitative phosphoproteomics reveals widespread full phosphorylation site occupancy during mitosis.
2010,
Pubmed
Ong,
Stable isotope labeling by amino acids in cell culture, SILAC, as a simple and accurate approach to expression proteomics.
2002,
Pubmed
Ow,
iTRAQ underestimation in simple and complex mixtures: "the good, the bad and the ugly".
2009,
Pubmed
Ow,
Quantitative shotgun proteomics of enriched heterocysts from Nostoc sp. PCC 7120 using 8-plex isobaric peptide tags.
2008,
Pubmed
Pichler,
Peptide labeling with isobaric tags yields higher identification rates using iTRAQ 4-plex compared to TMT 6-plex and iTRAQ 8-plex on LTQ Orbitrap.
2010,
Pubmed
Ting,
MS3 eliminates ratio distortion in isobaric multiplexed quantitative proteomics.
2011,
Pubmed
Tolonen,
Proteome-wide systems analysis of a cellulosic biofuel-producing microbe.
2011,
Pubmed
Wenger,
Gas-phase purification enables accurate, multiplexed proteome quantification with isobaric tagging.
2011,
Pubmed
Wysocki,
Mobile and localized protons: a framework for understanding peptide dissociation.
2000,
Pubmed
