Adaptation of a commonly used, chemically defined medium for human embryonic stem cells to stable isotope labeling with amino acids in cell culture

Albert R. Liberski, Muna Al-Noubi, Zahra H. Rahman, Najeeb Halabi, Shaymaa Dib, Rasha Al Mismar, Anja Billing, Roopesh Krishnankutty, Faizzan S. Ahmad, Christophe M. Raynaud, Arash Rafii Tabrizi, Kasper Engholm-Keller, Johannes Graumann

Research output: Contribution to journalArticle

7 Citations (Scopus)

Abstract

Metabolic labeling with stable isotopes is a prominent technique for comparative quantitative proteomics, and stable isotope labeling with amino acids in cell culture (SILAC) is the most commonly used approach. SILAC is, however, traditionally limited to simple tissue culture regimens and only rarely employed in the context of complex culturing conditions as those required for human embryonic stem cells (hESCs). Classic hESC culture is based on the use of mouse embryonic fibroblasts (MEFs) as a feeder layer, and as a result, possible xenogeneic contamination, contribution of unlabeled amino acids by the feeders, interlaboratory variability of MEF preparation, and the overall complexity of the culture system are all of concern in conjunction with SILAC. We demonstrate a feeder-free SILAC culture system based on a customized version of a commonly used, chemically defined hESC medium developed by Ludwig et al. and commercially available as mTeSR1 [mTeSR1 is a trade mark of WiCell (Madison, WI) licensed to STEMCELL Technologies (Vancouver, Canada)]. This medium, together with adjustments to the culturing protocol, facilitates reproducible labeling that is easily scalable to the protein amounts required by proteomic work flows. It greatly enhances the usability of quantitative proteomics as a tool for the study of mechanisms underlying hESCs differentiation and self-renewal. Associated data have been deposited to the ProteomeXchange with the identifier PXD000151.

Original languageEnglish
Pages (from-to)3233-3245
Number of pages13
JournalJournal of Proteome Research
Volume12
Issue number7
DOIs
Publication statusPublished - 5 Jul 2013

Fingerprint

Isotope Labeling
Stem cells
Cell culture
Isotopes
Labeling
Cell Culture Techniques
Proteomics
Amino Acids
Fibroblasts
Feeder Cells
Tissue culture
Trademarks
Workflow
Canada
Cell Differentiation
Contamination
Technology
Human Embryonic Stem Cells
Proteins

Keywords

  • chemically defined medium
  • human embryonic stem cells
  • metabolic labeling
  • SILAC

ASJC Scopus subject areas

  • Biochemistry
  • Chemistry(all)

Cite this

Adaptation of a commonly used, chemically defined medium for human embryonic stem cells to stable isotope labeling with amino acids in cell culture. / Liberski, Albert R.; Al-Noubi, Muna; Rahman, Zahra H.; Halabi, Najeeb; Dib, Shaymaa; Al Mismar, Rasha; Billing, Anja; Krishnankutty, Roopesh; Ahmad, Faizzan S.; Raynaud, Christophe M.; Tabrizi, Arash Rafii; Engholm-Keller, Kasper; Graumann, Johannes.

In: Journal of Proteome Research, Vol. 12, No. 7, 05.07.2013, p. 3233-3245.

Research output: Contribution to journalArticle

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AU - Ahmad, Faizzan S.

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