| An exclusion list based label-free proteome quantification approach using an LTQ Orbitrap. | |
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MedLine Citation:
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PMID: 22328225 Owner: NLM Status: In-Data-Review |
Abstract/OtherAbstract:
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RATIONALE: Label-based mass spectrometry is a powerful tool for large-scale protein identification and quantification. However, it requires the chemical or metabolic incorporation of the labeled compound(s) which can be difficult to attain, e.g. for non-cultivable organisms or scarce sample, such as biopsies. Therefore, we set out to develop and validate an efficient label-free liquid chromatography/tandem mass spectrometry (LC/MS/MS) workflow based on optimized instrument settings and incremental exclusion lists. METHODS: To increase the number of quantified peptides an incremental exclusion list was incorporated along with optimized instrument settings for the used LTQ Orbitrap. As a proof of concept, label-free quantification data from this optimized approach were compared to the results of control measurements without exclusion lists and of an in vivo metabolic labeling GeLC/MS/MS experiment. The data were drawn from Staphylococcus aureus whole cell lysates of non-stressed and nitric oxide (NO)-stressed cells. RESULTS: Compared to MS analysis without exclusion lists the new approach resulted in an increased number of identified peptides, enabling label-free quantification of more than 990 S. aureus proteins. With respect to the number of quantified proteins and differences in protein levels between the control and NO-treated samples the results of the new method were consistent with those of the GeLC/MS/MS experiment. CONCLUSIONS: The application of exclusion lists and optimized instrument settings in LC/MS/MS analysis significantly enhances the sensitivity and resolution of label-free protein identification and quantification. Therefore, the new workflow is a powerful alternative to label-based quantification methods. Copyright © 2012 John Wiley & Sons, Ltd. |
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Authors:
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Jan Muntel; Michael Hecker; Dörte Becher |
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Publication Detail:
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Type: Journal Article |
Journal Detail:
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Title: Rapid communications in mass spectrometry : RCM Volume: 26 ISSN: 1097-0231 ISO Abbreviation: Rapid Commun. Mass Spectrom. Publication Date: 2012 Mar |
Date Detail:
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Created Date: 2012-02-13 Completed Date: - Revised Date: - |
Medline Journal Info:
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Nlm Unique ID: 8802365 Medline TA: Rapid Commun Mass Spectrom Country: England |
Other Details:
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Languages: eng Pagination: 701-9 Citation Subset: IM |
Copyright Information:
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Copyright © 2012 John Wiley & Sons, Ltd. |
Affiliation:
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Institute for Microbiology, Ernst Moritz Arndt University Greifswald, Friedrich-Ludwig-Jahn-Str. 15, D-17489, Greifswald, Germany. |
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From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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