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Introducing AAA-MS, a rapid and sensitive method for amino acid analysis using isotope-dilution and high-resolution mass spectrometry.
MedLine Citation:
PMID:  22681258     Owner:  NLM     Status:  Publisher    
Abstract/OtherAbstract:
Accurate quantification of pure peptides and proteins is essential for biotechnology, clinical chemistry, proteomics and systems biology. The reference method to quantify peptides and proteins is amino acid analysis (AAA). This consists in an acidic hydrolysis followed by chromatographic separation and spectrophotometric detection of amino acids. Although widely used, this method displays some limitations, in particular the need for large amounts of starting material. Driven by the need to quantify isotope-dilution standards used for absolute quantitative proteomics, particularly stable isotope-labeled (SIL) peptides and PSAQ™ proteins, we developed a new AAA assay (AAA-MS). This method requires neither derivatization nor chromatographic separation of amino acids. It is based on rapid microwave-assisted acidic hydrolysis followed by high-resolution mass spectrometry analysis of amino acids. Quantification is performed by comparing MS signals from labeled amino acids (SIL peptide- and PSAQ-derived) with those of unlabeled amino acids originating from co-hydrolyzed NIST standard reference materials. For both SIL peptides and PSAQ standards, AAA-MS quantification results were consistent with classical AAA measurements. Compared to AAA assay, AAA-MS was much faster and was 100 fold more sensitive for peptide and protein quantification. Finally, thanks to the development of a labeled protein standard, we also extended AAA-MS analysis to the quantification of unlabeled proteins.
Authors:
Mathilde Louwagie; Sylvie Kieffer-Jaquinod; Veronique Dupierris; Yohann Couté; Christophe Bruley; Jérôme Garin; Alain Dupuis; Michel Jaquinod; Virginie Brun
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Publication Detail:
Type:  JOURNAL ARTICLE     Date:  2012-6-8
Journal Detail:
Title:  Journal of proteome research     Volume:  -     ISSN:  1535-3907     ISO Abbreviation:  -     Publication Date:  2012 Jun 
Date Detail:
Created Date:  2012-6-11     Completed Date:  -     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  101128775     Medline TA:  J Proteome Res     Country:  -    
Other Details:
Languages:  ENG     Pagination:  -     Citation Subset:  -    
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