title: Exploiting the multiplexing capabilities of tandem mass tags for high-throughput estimation of cellular protein abundances by mass spectrometry.
abstract: The generation of dynamic models of biological processes critically depends on the determination of precise cellular concentrations of biomolecules. Measurements of system-wide absolute protein levels are particularly valuable information in systems biology. Recently, mass spectrometry based proteomics approaches have been developed to estimate protein concentrations on a proteome-wide scale. However, for very complex proteomes, fractionation steps are required, increasing samples number and instrument analysis time. As a result, the number of full proteomes that can be routinely analyzed is limited. Here we combined absolute quantification strategies with the multiplexing capabilities of isobaric tandem mass tags to determine cellular protein abundances in a high throughput and proteome-wide scale even for highly complex biological systems, such as a whole human cell line. We generated two independent data sets to demonstrate the power of the approach regarding sample throughput, dynamic range, quantitative precision and accuracy as well as proteome coverage in comparison to existing mass spectrometry based strategies.
authors.affiliation: Biozentrum, University of Basel, Klingelbergstrasse 50/70, 4056 Basel, Switzerland.
authors.forename: Erik
authors.initials: E
authors.lastname: Ahrné
authors.affiliation: Quantitative Gene Expression Group, MRC Clinical Sciences Centre, Imperial College London, Hammersmith Hospital Campus, Du Cane Road, London W12 0NN, UK.
authors.forename: Amalia
authors.initials: A
authors.lastname: Martinez-Segura
authors.affiliation: Biozentrum, University of Basel, Klingelbergstrasse 50/70, 4056 Basel, Switzerland.
authors.forename: Afzal Pasha
authors.initials: AP
authors.lastname: Syed
authors.affiliation: Biozentrum, University of Basel, Klingelbergstrasse 50/70, 4056 Basel, Switzerland.
authors.forename: Arnau
authors.initials: A
authors.lastname: Vina-Vilaseca
authors.affiliation: Biozentrum, University of Basel, Klingelbergstrasse 50/70, 4056 Basel, Switzerland.
authors.forename: Andreas J
authors.initials: AJ
authors.lastname: Gruber
authors.affiliation: Quantitative Gene Expression Group, MRC Clinical Sciences Centre, Imperial College London, Hammersmith Hospital Campus, Du Cane Road, London W12 0NN, UK.
authors.forename: Samuel
authors.initials: S
authors.lastname: Marguerat
authors.affiliation: Biozentrum, University of Basel, Klingelbergstrasse 50/70, 4056 Basel, Switzerland. Electronic address: alex.schmidt@unibas.ch.
authors.forename: Alexander
authors.initials: A
authors.lastname: Schmidt
mesh_terms: D002853:Chromatography, Liquid / Q000379:methods; D057809:HEK293 Cells; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans; D007553:Isotope Labeling / Q000379:methods*; D013058:Mass Spectrometry / Q000379:methods; D053719:Tandem Mass Spectrometry / Q000379:methods*
keywords: Absolute protein quantification; HEK 293; Mass spectrometry; Schizosaccharomyces pombe; Tandem mass tags; iBAQ