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dc.contributor.authorPotier, D
dc.contributor.authorGriffiths, John R
dc.contributor.authorUnwin, R
dc.contributor.authorWalker, M
dc.contributor.authorCarrick, E
dc.contributor.authorWillamson, A
dc.contributor.authorWhetton, A
dc.date.accessioned2012-12-05T18:06:24Z
dc.date.available2012-12-05T18:06:24Z
dc.date.issued2012-07-03
dc.identifier.citationAn assessment of peptide enrichment methods employing mTRAQ quantification approaches. 2012, 84 (13):5604-10 Anal Chemen_GB
dc.identifier.issn1520-6882
dc.identifier.pmid22762262
dc.identifier.doi10.1021/ac300584y
dc.identifier.urihttp://hdl.handle.net/10541/254637
dc.description.abstractThe human plasma peptidome has potential in biomarker discovery not least because the plasma proteome is a challenging matrix due to its complexity and dynamic range. However, methods to significantly reduce the amount of protein present in plasma while retaining the less abundant peptides present in plasma samples has been a major issue. Here, we present a novel strategy which has been employed to assess the effectiveness of removing interfering proteins while retaining peptides of interest. To monitor peptide retention, a spiked in digested protein, in this case a synthetic QconCAT protein, was employed. This enabled a variety of target analytes (peptides) to be monitored for their retention in liquid phase, providing a broader picture of peptide loss from each method assessed. The incorporation of mTRAQ labeling allowed the presence of each peptide to be monitored, and accurate peptide losses to be determined in a Selected Reaction Monitoring (SRM) assay, thus, enabling an objective semiquantitative conclusion to be drawn regarding the suitability of each method for protein removal and peptide retention. We also assessed a range of methods for retaining nontryptic peptides in a plasma peptidomics workflow. From these data, we determined an optimal workflow for removing intact protein, while retaining peptides for MS-based analyses.
dc.language.isoenen
dc.rightsArchived with thanks to Analytical chemistryen_GB
dc.subject.meshAmino Acid Sequence
dc.subject.meshBlood Proteins
dc.subject.meshChemical Precipitation
dc.subject.meshElectrophoresis, Polyacrylamide Gel
dc.subject.meshFiltration
dc.subject.meshHumans
dc.subject.meshMass Spectrometry
dc.subject.meshMolecular Sequence Data
dc.subject.meshPeptides
dc.subject.meshPlasma
dc.subject.meshSilver Staining
dc.subject.meshSolid Phase Extraction
dc.titleAn assessment of peptide enrichment methods employing mTRAQ quantification approaches.en
dc.typeArticleen
dc.contributor.departmentSchool of Cancer & Enabling Sciences, Wolfson Molecular Imaging Centre, University of Manchester, Manchester Academic Health Science Centre, Manchester, UK.en_GB
dc.identifier.journalAnalytical Chemistryen_GB
html.description.abstractThe human plasma peptidome has potential in biomarker discovery not least because the plasma proteome is a challenging matrix due to its complexity and dynamic range. However, methods to significantly reduce the amount of protein present in plasma while retaining the less abundant peptides present in plasma samples has been a major issue. Here, we present a novel strategy which has been employed to assess the effectiveness of removing interfering proteins while retaining peptides of interest. To monitor peptide retention, a spiked in digested protein, in this case a synthetic QconCAT protein, was employed. This enabled a variety of target analytes (peptides) to be monitored for their retention in liquid phase, providing a broader picture of peptide loss from each method assessed. The incorporation of mTRAQ labeling allowed the presence of each peptide to be monitored, and accurate peptide losses to be determined in a Selected Reaction Monitoring (SRM) assay, thus, enabling an objective semiquantitative conclusion to be drawn regarding the suitability of each method for protein removal and peptide retention. We also assessed a range of methods for retaining nontryptic peptides in a plasma peptidomics workflow. From these data, we determined an optimal workflow for removing intact protein, while retaining peptides for MS-based analyses.


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