Chapter title |
Testing Suitability of Cell Cultures for SILAC-Experiments Using SWATH-Mass Spectrometry.
|
---|---|
Chapter number | 8 |
Book title |
Proteomics in Systems Biology
|
Published in |
Methods in molecular biology, January 2016
|
DOI | 10.1007/978-1-4939-3341-9_8 |
Pubmed ID | |
Book ISBNs |
978-1-4939-3339-6, 978-1-4939-3341-9
|
Authors |
Yvonne Reinders, Daniel Völler, Anja-K. Bosserhoff, Peter J. Oefner, Jörg Reinders |
Editors |
Jörg Reinders |
Abstract |
Precise quantification is a major issue in contemporary proteomics. Both stable-isotope-labeling and label-free methods have been established for differential protein quantification and both approaches have different advantages and disadvantages. The present protocol uses the superior precision of label-free SWATH-mass spectrometry to test for suitability of cell lines for a SILAC-labeling approach as systematic regulations may be introduced upon incorporation of the "heavy" amino acids. The SILAC-labeled cell cultures can afterwards be used for further analyses where stable-isotope-labeling is mandatory or has substantial advantages over label-free approaches such as pulse-chase-experiments and differential protein interaction analyses based on co-immunoprecipitation. As SWATH-mass spectrometry avoids the missing-value-problem typically caused by undersampling in highly complex samples and shows superior precision for the quantification, it is better suited for the detection of systematic changes caused by the SILAC-labeling and thus, can serve as a useful tool to test cell lines for changes upon SILAC-labeling. |
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