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Development and optimization of a metabolomic method for analysis of adherent cell cultures.

Anders P H AP Danielsson, Thomas T Moritz, Hindrik H Mulder, and Peter P Spégel

Anal Biochem 404(1):30-9 (2010) PMID 20417172

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 24h.

Copyright © 2010 Elsevier Ltd. All rights reserved.

DOI: 10.1016/j.ab.2010.04.013
Version: za2963e q8za5 q8zb3 q8zc5 q8zdf q8ze7 q8zfa q8zg0

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