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Validation of reference gene stability for APAP hepatotoxicity studies in different in vitro systems and identification of novel potential toxicity biomarkers
Liver cell lines and primary hepatocytes are becoming increasingly valuable for in vitro toxicogenomic studies, with RT-qPCR enabling the analysis of gene expression profiles following exposure to potential hepatotoxicants. Supporting the accurate normalisation of RT-qPCR data requires the identification of reference genes which have stable expression during in vitro toxicology studies. This paper includes a comprehensive analysis of reference gene stability in two routinely used cell types, (HepG2 cells and primary rat hepatocytes), and two in vitro culture systems, (2D monolayer and 3D scaffolds). A robust reference gene validation strategy was performed, consisting of geNorm analysis, to test for pair wise variation in gene expression, and statistical analysis using analysis of variance. This strategy identified stable reference genes with respect to acetaminophen treatment and time in HepG2 cells (GAPDH and PPIA), and with respect to acetaminophen treatment and culture condition in primary hepatocytes (18S rRNA and alpha-tubulin).
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- Author - Fox BC
- Reference Number - Toxicol In Vitro. 2010 Aug 21. [Epub ahead of print]
Last modified on
16 September 2010.