Antibody-based capture of target peptides in multiple reaction monitoring experiments
(2015) In Methods in Molecular Biology 1293. p.35-123- Abstract
Targeted quantitative mass spectrometry of immunoaffinity-enriched peptides, termed immuno-multiple reaction monitoring (iMRM), is a powerful method for determining the relative abundance of proteins in complex mixtures, like plasma or whole tissue. This technique combines 1,000-fold enrichment potential of antibodies for target peptides with the selectivity of multiple reaction monitoring mass spectrometry (MRM-MS). Using heavy isotope-labeled peptide counterparts as internal standards ensures high levels of precision. Further, LC-MRM-MS selectivity allows for multiplexing; antibodies recognizing different peptides can be added directly to a single mixture without subjecting to interferences common to other multiple antibody protein... (More)
Targeted quantitative mass spectrometry of immunoaffinity-enriched peptides, termed immuno-multiple reaction monitoring (iMRM), is a powerful method for determining the relative abundance of proteins in complex mixtures, like plasma or whole tissue. This technique combines 1,000-fold enrichment potential of antibodies for target peptides with the selectivity of multiple reaction monitoring mass spectrometry (MRM-MS). Using heavy isotope-labeled peptide counterparts as internal standards ensures high levels of precision. Further, LC-MRM-MS selectivity allows for multiplexing; antibodies recognizing different peptides can be added directly to a single mixture without subjecting to interferences common to other multiple antibody protein assays. Integrated extracted ion chromatograms (XIC) of product ions from endogenous unlabeled "light" peptide and stable isotope-labeled internal standard "heavy" peptides are used to generate a light/heavy peak area ratio. This ratio is proportional to the amount of peptide in the digestion mixture and can be used to estimate the concentration of protein in the sample.
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- author
- De Marchi, Tommaso LU ; Kuhn, Eric ; Carr, Steven A. and Umar, Arzu
- publishing date
- 2015
- type
- Chapter in Book/Report/Conference proceeding
- publication status
- published
- keywords
- Mass Spectrometry, Peptides, Proteomics, Journal Article, Research Support, Non-U.S. Gov't
- host publication
- Mammary Stem Cells : Methods and Protocols - Methods and Protocols
- series title
- Methods in Molecular Biology
- volume
- 1293
- pages
- 35 - 123
- publisher
- Humana Press
- external identifiers
-
- pmid:26040685
- scopus:84930655255
- DOI
- 10.1007/978-1-4939-2519-3_7
- language
- English
- LU publication?
- no
- id
- e2e5e3de-82ea-44d5-b192-3b796c1a70ff
- date added to LUP
- 2017-06-27 14:28:22
- date last changed
- 2024-01-13 23:50:04
@inbook{e2e5e3de-82ea-44d5-b192-3b796c1a70ff,
abstract = {{<p>Targeted quantitative mass spectrometry of immunoaffinity-enriched peptides, termed immuno-multiple reaction monitoring (iMRM), is a powerful method for determining the relative abundance of proteins in complex mixtures, like plasma or whole tissue. This technique combines 1,000-fold enrichment potential of antibodies for target peptides with the selectivity of multiple reaction monitoring mass spectrometry (MRM-MS). Using heavy isotope-labeled peptide counterparts as internal standards ensures high levels of precision. Further, LC-MRM-MS selectivity allows for multiplexing; antibodies recognizing different peptides can be added directly to a single mixture without subjecting to interferences common to other multiple antibody protein assays. Integrated extracted ion chromatograms (XIC) of product ions from endogenous unlabeled "light" peptide and stable isotope-labeled internal standard "heavy" peptides are used to generate a light/heavy peak area ratio. This ratio is proportional to the amount of peptide in the digestion mixture and can be used to estimate the concentration of protein in the sample.</p>}},
author = {{De Marchi, Tommaso and Kuhn, Eric and Carr, Steven A. and Umar, Arzu}},
booktitle = {{Mammary Stem Cells : Methods and Protocols}},
keywords = {{Mass Spectrometry; Peptides; Proteomics; Journal Article; Research Support, Non-U.S. Gov't}},
language = {{eng}},
pages = {{35--123}},
publisher = {{Humana Press}},
series = {{Methods in Molecular Biology}},
title = {{Antibody-based capture of target peptides in multiple reaction monitoring experiments}},
url = {{http://dx.doi.org/10.1007/978-1-4939-2519-3_7}},
doi = {{10.1007/978-1-4939-2519-3_7}},
volume = {{1293}},
year = {{2015}},
}