An insight into iTRAQ: where do we stand now?

Abstract

The iTRAQ (isobaric tags for relative and absolute quantification) technique is widely employed in proteomic workflows requiring relative quantification. Here, we review the iTRAQ literature; in particular, we focus on iTRAQ usage in relation to other commonly used quantitative techniques e.g. stable isotope labelling in culture (SILAC), label-free methods and selected reaction monitoring (SRM). As a result, we identify several issues arising with respect to iTRAQ. Perhaps frustratingly, iTRAQ's attractiveness has been undermined by a number of technical and analytical limitations: it may not be truly quantitative, as the changes in abundance reported will generally be underestimated. We discuss weaknesses and strengths of iTRAQ as a methodology for relative quantification in the light of this and other technical issues. We focus on technical developments targeted at iTRAQ accuracy and precision, use of 4-plex over 8-plex reagents and application of iTRAQ to post-translational modification (PTM) workflows. We also discuss iTRAQ in relation to label-free approaches, to which iTRAQ is losing ground.

Label-Free Protein Quantification for Plant Golgi Protein Localization and Abundance

“…Label-free quantification is an increasingly popular alternative to isotopic tagging quantitative methods; it does not require labeling reagents and can be applied to an unlimited number of samples (Neilson et al, 2011;Evans et al, 2012). This is particularly appealing within plant proteomics, because the most conventional labeling strategy, Stable Isotope Labeling by Amino Acids in Cell Culture, is not easily suited for quantitative plant proteomic studies.…”

Advances in proteomics for production strain analysis

“…Gel free quantification methods achieve protein identification and relative quantification using MS with significant advantages over 2DE [10]. Protein and peptide labeling methods, metabolic (eg SILAC) or T'AQ -methods are increasingly gaining in popularity [11,12]. To date, no direct comparison of all these techniques has been reported ( Table 1).…”

Post-translational Modifications and Mass Spectrometry Detection

“…This approach allows the simultaneous analysis of four to eight distinct samples, depending on the reagents used (250). The interested reader may refer to (251) for an updated review of iTRAQ methods and applications. All these isotope labeling methodologies have been successfully employed in the quantification of PTMs, for example, in the characterization of grapevine response to infection by simultaneous monitoring protein phosphorylation, acetylation and glycosylation (252).…”