2011
DOI: 10.1021/pr200455s
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Evaluation of Different Multidimensional LC–MS/MS Pipelines for Isobaric Tags for Relative and Absolute Quantitation (iTRAQ)-Based Proteomic Analysis of Potato Tubers in Response to Cold Storage

Abstract: Cold-induced sweetening in potato tubers is a costly problem for the food industry. To systematically identify the proteins associated with this process, we employed a comparative proteomics approach using isobaric, stable isotope coded labels to compare the proteomes of potato tubers after 0 and 5 months of storage at 5 °C. We evaluated both high pH reverse phase (hpRP) liquid chromatography (LC) and off-gel electrophoresis (OGE) as first dimension fractionation methods followed by nanoLC-MS/MS, using two hig… Show more

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Cited by 116 publications
(122 citation statements)
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“…As co-isolation interference is dependent on sample complexity and the number of co-eluting peptides, improving chromatographic separation of peptides using higher resolution methods can somewhat improve quantification accuracy (22,23). We see a similar effect after IMAC enrichment of phosphopeptides as iTRAQ ratio compression is smaller for phosphoproteome samples compared with proteome samples due to their overall lower complexity.…”
Section: Discussionmentioning
confidence: 63%
“…As co-isolation interference is dependent on sample complexity and the number of co-eluting peptides, improving chromatographic separation of peptides using higher resolution methods can somewhat improve quantification accuracy (22,23). We see a similar effect after IMAC enrichment of phosphopeptides as iTRAQ ratio compression is smaller for phosphoproteome samples compared with proteome samples due to their overall lower complexity.…”
Section: Discussionmentioning
confidence: 63%
“…The tryptic peptides (5-10 l) were injected onto a PepMap C18 trap column (5 m, 300 m ϫ 5 mm, Dionex/Thermo Scientific, Sunnyvale CA) and separated on a PepMap C18 RP nano column (3 m, 75 m ϫ 15 cm, Dionex) that had been installed in the "CorConneX" device with a 10-m spray emitter (NewObjective, Woburn, MA) mounted in front of the Orbitrap orifice. Nano-LC conditions and Orbitrap instrument calibration/settings were as described previously (39). MS/MS data were acquired under parallel data-dependent acquisition mode.…”
Section: Methodsmentioning
confidence: 99%
“…Accuracy concerns, by contrast, are caused by predictable systematic biases (Karp et al, 2010). In the iTRAQ methodology, concerns regarding accuracy derived from the possibility that coeluted, near-isobaric ions that are isolated and fragmented together with the target ions lead to a distortion in reporter ion intensities (Ow et al, 2009;Karp et al, 2010;Ting et al, 2011;Yang et al, 2011). Several strategies have been employed to decrease or eliminate the accuracy bias.…”
Section: Accuracy and Precision Of Itraq Quantitationmentioning
confidence: 99%
“…Several strategies have been employed to decrease or eliminate the accuracy bias. One of these strategies comprises a sample pretreatment by SCX chromatography, which decreases sample complexity and enriches low-abundance peptides (Yang et al, 2011). A second strategy aims at eliminating protein quantification interference by applying triple-stage MS (Ting et al, 2011).…”
Section: Accuracy and Precision Of Itraq Quantitationmentioning
confidence: 99%