title: iTRAQ-Based Shotgun Proteomics Approach for Relative Protein Quantification.
abstract: Shotgun proteomics has a key role in quantitative estimation of proteins from biological systems under different conditions, which is crucial in the understanding of their functional roles. Isobaric tagging for relative and absolute quantitation (iTRAQ) mass spectrometry is based on pre-labeling of peptides with mass tags which allows the multiplex analysis of up to eight proteomes simultaneously. We describe here a detailed protocol for sample preparation and iTRAQ 4-plex labeling for relative quantification of multiple samples from human and plant tissues. We also present two strategies for peptide fractionation after the iTRAQ labeling protocol.
authors.affiliation: Laboratory of Protein Chemistry - Proteomics Unit, Chemistry Institute, Federal University of Rio de Janeiro, Bloco A, Lab 543, Avenida Athos da Silveira Ramos 149, Cidade Universitária, 21941-909, Rio de Janeiro, RJ, Brazil.
authors.forename: Erika Velásquez
authors.initials: EV
authors.lastname: Núñez
authors.affiliation: Laboratory of Protein Chemistry - Proteomics Unit, Chemistry Institute, Federal University of Rio de Janeiro, Bloco A, Lab 543, Avenida Athos da Silveira Ramos 149, Cidade Universitária, 21941-909, Rio de Janeiro, RJ, Brazil.
authors.forename: Gilberto Barbosa
authors.initials: GB
authors.lastname: Domont
authors.affiliation: Laboratory of Protein Chemistry - Proteomics Unit, Chemistry Institute, Federal University of Rio de Janeiro, Bloco A, Lab 543, Avenida Athos da Silveira Ramos 149, Cidade Universitária, 21941-909, Rio de Janeiro, RJ, Brazil. fabiocsn@ufrj.br.
authors.forename: Fábio César Sousa
authors.initials: FCS
authors.lastname: Nogueira
mesh_terms: D005591:Chemical Fractionation; D006801:Humans; D007553:Isotope Labeling; D010455:Peptides / Q000737:chemistry; D020543:Proteome; D040901:Proteomics / Q000379:methods*; D012984:Software; D053719:Tandem Mass Spectrometry / Q000379:methods
chemical_list: D010455:Peptides; D020543:Proteome
keywords: Animal tissues; Mass spectrometry; Plant tissues; Quantitative proteomics; Stable isotope labeling; iTRAQ