A label-free mass spectrometry method for the quantification of protein isotypes
dc.contributor.author | Winefield, Robert D. | |
dc.contributor.author | Williams, Todd D. | |
dc.contributor.author | Himes, Richard H. | |
dc.date.accessioned | 2017-01-12T19:18:26Z | |
dc.date.available | 2017-01-12T19:18:26Z | |
dc.date.issued | 2013-07-15 | |
dc.identifier.citation | Winefield, Robert D., Todd D. Williams, and Richard H. Himes. "A Label-free Mass Spectrometry Method for the Quantification of Protein Isotypes." Analytical Biochemistry 395.2 (2009): 217-23. | en_US |
dc.identifier.uri | http://hdl.handle.net/1808/22615 | |
dc.description.abstract | Successful quantitative mass spectrometry (MS) requires strategies to link the mass spectrometer response to the analyte abundance, with the response being dependent on more factors than just analyte abundance. Label-dependent strategies rely on the incorporation of an isotopically labeled internal standard into the sample. Current label-free strategies (performed without internal standards) are useful for analyzing samples that are unsuitable for isotopic labeling but are less accurate. Here we describe a label-free technique applicable to analysis of products from related genes (isotypes). This approach enables the invariant tryptic peptide sequences within the family to serve as “built-in” internal standards and the isotype-specific peptide sequences to report the amount of the various isotypes. A process of elimination segregates reliably trypsin-released standard and reporter peptides from unreliably released peptides. The specific MS response factors for these reporter and standard peptides can be determined using synthetic peptides. Analysis of HeLa tubulin digests revealed peptides from βI-, βII-, βIII-, βIVb-, and βV-tubulin, eight of which were suitable; along with five standard peptides for quantification of the β-tubulin isotypes. To show the utility of this method, we determined that βI-tubulin represented 77% and βIIItubulin represented 3.2% of the total HeLa β-tubulin. | en_US |
dc.publisher | Elsevier Masson | en_US |
dc.rights | This is an open access article under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License 3.0 (CC BY-NC-ND 3.0 US), which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no modifications or adaptations are made. | en_US |
dc.rights.uri | http://creativecommons.org/licenses/by-nc-nd/3.0/ | |
dc.subject | Label-free quantification | en_US |
dc.subject | Mass spectrometry | en_US |
dc.subject | Targeted proteomic analysis | en_US |
dc.subject | Tubulin isotype quantification | en_US |
dc.subject | Tubulin | en_US |
dc.title | A label-free mass spectrometry method for the quantification of protein isotypes | en_US |
dc.type | Article | en_US |
kusw.kuauthor | Williams, Todd D. | |
kusw.kudepartment | Mass Spectrometry Lab | en_US |
dc.identifier.doi | 10.1016/j.ab.2009.07.052 | en_US |
dc.identifier.orcid | https://orcid.org/0000-0002-0944-5407 | |
kusw.oaversion | Scholarly/refereed, author accepted manuscript | en_US |
kusw.oapolicy | This item meets KU Open Access policy criteria. | en_US |
dc.rights.accessrights | openAccess |
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Except where otherwise noted, this item's license is described as: This is an open access article under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License 3.0 (CC BY-NC-ND 3.0 US), which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no modifications or adaptations are made.