Chapter title |
Two Birds with One Stone: Parallel Quantification of Proteome and Phosphoproteome Using iTRAQ.
|
---|---|
Chapter number | 3 |
Book title |
Proteomics in Systems Biology
|
Published in |
Methods in molecular biology, January 2016
|
DOI | 10.1007/978-1-4939-3341-9_3 |
Pubmed ID | |
Book ISBNs |
978-1-4939-3339-6, 978-1-4939-3341-9
|
Authors |
Fiorella A. Solari, Laxmikanth Kollipara, Albert Sickmann, René P. Zahedi |
Editors |
Jörg Reinders |
Abstract |
Altered and abnormal levels of proteins and their phosphorylation states are associated with many disorders. Detection and quantification of such perturbations may provide a better understanding of pathological conditions and help finding candidates for treatment or biomarkers. Over the years, isobaric mass tags for relative quantification of proteins and protein phosphorylation by mass spectrometry have become increasingly popular. One of the most commonly used isobaric chemical tags is iTRAQ (isobaric tag for relative and absolute quantitation). In a typical iTRAQ-8plex experiment, a multiplexed sample amounts for up to 800 μg of peptides. Using state-of-the-art LC-MS approaches, only a fraction (~5 %) of such a sample is required to generate comprehensive quantitative data on the global proteome level, so that the bulk of the sample can be simultaneously used for quantitative phosphoproteomics. Here, we provide a simple and straightforward protocol to perform quantitative analyses of both proteome and phosphoproteome from the same sample using iTRAQ. |
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