Title
Effective correction of experimental errors in quantitative proteomics using stable isotope labeling by amino acids in cell culture (SILAC) Effective correction of experimental errors in quantitative proteomics using stable isotope labeling by amino acids in cell culture (SILAC)
Author
Faculty/Department
Faculty of Pharmaceutical, Biomedical and Veterinary Sciences . Biomedical Sciences
Publication type
article
Publication
Subject
Chemistry
Biology
Source (journal)
Journal of proteomics
Volume/pages
75(2012) :12 , p. 3720-3732
ISSN
1874-3919
ISI
000307086500028
Carrier
E
Target language
English (eng)
Full text (Publishers DOI)
Abstract
Accurate and reliable quantitative proteomics in cell culture has been considerably facilitated by the introduction of the stable isotope labeling by amino acids in cell culture (SILAC), combined with high resolution mass spectrometry. There are however several major sources of quantification errors that commonly occur with SILAC techniques, i.e. incomplete incorporation of isotopic amino acids, arginine-to-proline conversion, and experimental errors in final sample mixing. Dataset normalization is a widely adopted solution to such errors, however this may not completely prevent introducing incorrect expression ratios. Here we demonstrate that a label-swap replication of SILAC experiments was able to effectively correct experimental errors by averaging ratios measured in individual replicates using quantitative proteomics and phosphoproteomics of ligand treatment of neural cell cultures. Furthermore, this strategy was successfully applied to a SILAC triplet experiment, which presents a much more complicated experimental matrix, affected by both incomplete labeling and arginine-to-proline conversion. Based on our results, we suggest that SILAC experiments should be designed to incorporate label-swap replications for enhanced reliability in expression ratios. Published by Elsevier B.V.
E-info
https://repository.uantwerpen.be/docman/iruaauth/de0e97/8978924.pdf
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