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Mutagenesis Advance Access originally published online on September 11, 2008
Mutagenesis 2009 24(1):35-50; doi:10.1093/mutage/gen050
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© The Author 2008. Published by Oxford University Press on behalf of the UK Environmental Mutagen Society. All rights reserved. For permissions, please e-mail: journals.permissions@oxfordjournals.org.

Assessment of the genotoxicity of S9-generated metabolites using the GreenScreen HC GADD45a–GFP assay

Christopher Jagger{dagger}, Matthew Tate1,{dagger}, Paul A. Cahill, Chris Hughes, Andrew W. Knight, Nicholas Billinton* and Richard M. Walmsley1

Gentronix Ltd, CTF Building, 46 Grafton Street, Manchester M13 9NT, UK 1Faculty of Life Sciences, University of Manchester, Manchester M13 9PT, UK

Genotoxicity can be assessed by monitoring expression of a GADD45a–GFP reporter in the human lymphoblastoid cell line TK6. A flow cytometric method has been developed to effectively distinguish GFP fluorescence from coloured and fluorescent test samples as well from the S9 liver extracts used to generate metabolites from pro-genotoxins. The method includes the use of propidium iodide exclusion for the determination of cellular viability. This paper describes the method development, the derivation of decision thresholds for the identification of genotoxins using the method, and presents data from a 56-compound validation study of the method. The results illustrate that the method permitted the detection of the majority of pro-genotoxins tested and, importantly, the high specificity of the GADD45a–GFP assay was maintained.

* To whom correspondence should be addressed. Tel: +44 161 603 7661; Fax: +44 161 606 7337; Email: nick.billinton{at}gentronix.co.uk

{dagger} Christopher Jagger and Matthew Tate are joint first authors.

Received on June 9, 2008; revised on August 5, 2008; accepted on August 6, 2008.


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