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Article: Fully automatable two-dimensional reversed-phase capillary liquid chromatography with online tandem mass spectrometry for shotgun proteomics

TitleFully automatable two-dimensional reversed-phase capillary liquid chromatography with online tandem mass spectrometry for shotgun proteomics
Authors
KeywordsLC-MS/MS
Multidimensional LC
Protein profile
Technology
Issue Date2011
PublisherWiley - V C H Verlag GmbH & Co KGaA. The Journal's web site is located at http://www.wiley-vch.de/home/proteomics
Citation
Proteomics, 2011, v. 11 n. 11, p. 2308-2319 How to Cite?
AbstractHerein, we describe the development of a fully automatable technology that features online coupling of high-pH RP separation with conventional low-pH RP separation in a two-dimensional capillary liquid chromatography (2-D LC) system for shotgun proteomics analyses. The complete analysis comprises 13 separation cycles, each involving transfer of the eluate from the first-dimension, high-pH RP separation onto the second RP dimension for further separation. The solvent strength increases across the 13 fractions (cycles) to elute all peptides for further resolution on the second-dimension, low-pH RP separation, each under identical gradient-elution conditions. The total run time per analysis is 52h. In triplicate analyses of a lysate of mouse embryonic fibroblasts, we used this technology to identify 2431 non-redundant proteins, of which 50% were observed in all three replicates. A comparison of RP-RP 2-D LC and strong cation exchange-RP 2-D LC analyses reveals that the two technologies identify primarily different peptides, thereby underscoring the differences in their separation chemistries. © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
Persistent Identifierhttp://hdl.handle.net/10722/135037
ISSN
2015 Impact Factor: 4.079
2015 SCImago Journal Rankings: 1.476
ISI Accession Number ID
Funding AgencyGrant Number
Hong Kong Research Grants Council, Hong Kong Special Administrative Region, ChinaHKU7018/09P
HKU3/07C
Science and Technology Development Fund of Macau SAR045/2007/A3
Research Committee, University of MacauUL017
Hong Kong RGC
Funding Information:

This study was supported by the Hong Kong Research Grants Council (Project No. HKU7018/09P and HKU3/07C), Hong Kong Special Administrative Region, China; the Science and Technology Development Fund of Macau SAR (Ref. No. 045/2007/A3); and the Research Committee, University of Macau (Ref. No. UL017). E. L., M. P. Y. L., and R. K. thank the Hong Kong RGC for supporting their studentships. The authors thank Dr. Ronald T. K. Pang for providing the STO Cell lysates.

References

 

DC FieldValueLanguage
dc.contributor.authorSiu, SOen_HK
dc.contributor.authorLam, MPYen_HK
dc.contributor.authorLau, Een_HK
dc.contributor.authorKong, RPWen_HK
dc.contributor.authorLee, SMYen_HK
dc.contributor.authorChu, IKen_HK
dc.date.accessioned2011-07-27T01:26:23Z-
dc.date.available2011-07-27T01:26:23Z-
dc.date.issued2011en_HK
dc.identifier.citationProteomics, 2011, v. 11 n. 11, p. 2308-2319en_HK
dc.identifier.issn1615-9853en_HK
dc.identifier.urihttp://hdl.handle.net/10722/135037-
dc.description.abstractHerein, we describe the development of a fully automatable technology that features online coupling of high-pH RP separation with conventional low-pH RP separation in a two-dimensional capillary liquid chromatography (2-D LC) system for shotgun proteomics analyses. The complete analysis comprises 13 separation cycles, each involving transfer of the eluate from the first-dimension, high-pH RP separation onto the second RP dimension for further separation. The solvent strength increases across the 13 fractions (cycles) to elute all peptides for further resolution on the second-dimension, low-pH RP separation, each under identical gradient-elution conditions. The total run time per analysis is 52h. In triplicate analyses of a lysate of mouse embryonic fibroblasts, we used this technology to identify 2431 non-redundant proteins, of which 50% were observed in all three replicates. A comparison of RP-RP 2-D LC and strong cation exchange-RP 2-D LC analyses reveals that the two technologies identify primarily different peptides, thereby underscoring the differences in their separation chemistries. © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.en_HK
dc.languageengen_US
dc.publisherWiley - V C H Verlag GmbH & Co KGaA. The Journal's web site is located at http://www.wiley-vch.de/home/proteomicsen_HK
dc.relation.ispartofProteomicsen_HK
dc.subjectLC-MS/MSen_HK
dc.subjectMultidimensional LCen_HK
dc.subjectProtein profileen_HK
dc.subjectTechnologyen_HK
dc.subject.meshChromatography, Reverse-Phase - methods-
dc.subject.meshPeptide Fragments - analysis - chemistry-
dc.subject.meshPeptide Mapping - methods-
dc.subject.meshProteomics - instrumentation - methods-
dc.subject.meshTandem Mass Spectrometry - methods-
dc.titleFully automatable two-dimensional reversed-phase capillary liquid chromatography with online tandem mass spectrometry for shotgun proteomicsen_HK
dc.typeArticleen_HK
dc.identifier.emailChu, IK:ivankchu@hku.hken_HK
dc.identifier.authorityChu, IK=rp00683en_HK
dc.description.naturelink_to_subscribed_fulltext-
dc.identifier.doi10.1002/pmic.201100110en_HK
dc.identifier.pmid21548098-
dc.identifier.scopuseid_2-s2.0-79956270700en_HK
dc.identifier.hkuros186753en_US
dc.relation.referenceshttp://www.scopus.com/mlt/select.url?eid=2-s2.0-79956270700&selection=ref&src=s&origin=recordpageen_HK
dc.identifier.volume11en_HK
dc.identifier.issue11en_HK
dc.identifier.spage2308en_HK
dc.identifier.epage2319en_HK
dc.identifier.eissn1615-9861-
dc.identifier.isiWOS:000291087700014-
dc.publisher.placeGermanyen_HK
dc.identifier.scopusauthoridSiu, SO=8603087200en_HK
dc.identifier.scopusauthoridLam, MPY=35302594800en_HK
dc.identifier.scopusauthoridLau, E=35302963200en_HK
dc.identifier.scopusauthoridKong, RPW=35217869000en_HK
dc.identifier.scopusauthoridLee, SMY=35233892600en_HK
dc.identifier.scopusauthoridChu, IK=7103327484en_HK

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