Titlebook: LC-MS/MS in Proteomics; Methods and Applicat Pedro R. Cutillas,John F. Timms Book 2010 Springer Science+Business Media, LLC 2010 Biological

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Quantification of Proteins by iTRAQd elsewhere in this volume. In this chapter we will describe the use of isobaric tags for relative and absolute quantification (iTRAQ). These chemical tags attach to all peptides in a protein digest via free amines at the peptide N-terminus and on the side chain of lysine residues. Labelled samples

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Protocol for Quantitative Proteomics of Cellular Membranes and Membrane Raftson digestion of the proteins prior to ESI-LC-MS/MS. In-gel digestion yields more comprehensive proteomic and protein coverage of membrane/membrane raft samples, for example by LC-MS/MS of protein samples resolved by 1D SDS-polyacrylamide gel electrophoresis. Although this type of analysis can be per

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In-Depth Analysis of Protein Phosphorylation by Multidimensional Ion Exchange Chromatography and Mase we present a detailed protocol to enrich phosphopeptides from total cell lysates in a form amenable to downstream analysis by mass spectrometry. Using these techniques, we have detected several thousands of phosphorylation sites in the NIH-3T3 cell line.

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Selected Reaction Monitoring Applied to Quantitative Proteomicsasingly applied to quantitative proteomics because of its selectivity (two levels of mass selection), its sensitivity (non-scanning mode), and its wide dynamic range. This account describes the different steps in the design and the experimental setup of SRM experiments.

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Protocol for Quantitative Proteomics of Cellular Membranes and Membrane Raftsor membrane raft protein samples to be isolated, but not separated, as unresolved bands in a gel. Focusing as a single band enables the confident excision of different samples in their entirety, to be digested, labelled, and fractionated for quantitative mass spectrometric analysis.

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Organelle Proteomics by Label-Free and SILAC-Based Protein Correlation Profilinged on protein correlation profiling and quantitative mass spectrometry to sort out likely candidates. The organelle inventories defined by these methods are suitable to guide future functional experiments.