Abstract

In this investigation, a gas chromatography/mass spectrometry (GC/MS)-based metabolomic protocol for adherent cell cultures was developed using statistical design of experiments. Cell disruption, metabolite extraction, and the GC/MS settings were optimized aiming at a gentle, unbiased, sensitive, and high-throughput metabolomic protocol. Due to the heterogeneity of the metabolome and the inherent selectivity of all analytical techniques, development of unbiased protocols is highly complex. Changing one parameter of the protocol may change the response of many groups of metabolites. In this investigation, statistical design of experiments and multivariate analysis also allowed such interaction effects to be taken into account. The protocol was validated with respect to linear range, precision, and limit of detection in a clonal rat insulinoma cell line (INS-1 832/13). The protocol allowed high-throughput profiling of metabolites covering the major metabolic pathways. The majority of metabolites displayed a linear range from a single well in a 96-well plate up to a 10 cm culture dish. The method allowed a total of 47 analyses to be performed in 24 h. (C) 2010 Elsevier Inc. All rights reserved.

Keywords

Metabolomics; Statistical design of experiments; Gas chromatography; Mass spectrometry; Cell culture

Published in

Analytical Biochemistry
2010, Volume: 404, number: 1, pages: 30-39
Publisher: ACADEMIC PRESS INC ELSEVIER SCIENCE

SLU Authors

• Moritz, Thomas

  • Department of Forest Genetics and Plant Physiology, Swedish University of Agricultural Sciences
    

UKÄ Subject classification

Forest Science


Publication identifier

DOI: https://doi.org/10.1016/j.ab.2010.04.013

Permanent link to this page (URI)

https://res.slu.se/id/publ/48059