Amplified Recombinant Cell Bioassay for the Detection of Dioxin and Related Ah Receptor Ligands
10- fold to 100-fold increase in the minimal detection limits for dioxin and related chemicals Dramatically enhanced overall response compared to other bioanalytical methods Rapid and very low cost Mouse, rat and human hepatoma cell lines, stably transfected with the AhR-responsive CALUX luciferase reporter plasmids, of this invention have dramatically improved sensitivity and reporter gene responsiveness to TCDD and related AhR agonists. Improved luciferase activity observed at all inducer concentrations. Increased activity and sensitivity allows for more accurate determinations of the induction response at the lower end of the concentration response curve, and results in a lower limit of chemical detection with these new cells Can be used in a range of microplate formats (96 to 1536 well), reducing the amount of sample and reagents needed for cell growth and analysis while also reducing analysis cost and increasing sample throughput.
Detection and relative quantitation of dioxin and dioxin-like chemicals in a wide variety of matrices, including primary matrices of concern such as food, feed, biological (tissue, milk and blood) and environmental samples High throughput screening applications.
The Ah receptor (AhR) is a ligand-dependent transcription factor that mediates the ability of dioxin and related halogenated aromatic hydrocarbons (HaHs) to induce gene expression and to produce toxicity. Cell-based bioassays for dioxin-like chemicals (commonly referred to as CALUX bioassays) respond to dioxins and related chemicals with the induction of luciferase in a time-, dose-, AhR-, and chemical specific manner. While high-resolution instrumental analysis methods are the gold standard method for detection of dioxin-like chemicals and provide very accurate measurements of HAH presence, these procedures require expensive instrumentation, are time-consuming, and are impractical for large-scale screening analysis. There is a need for rapid, inexpensive and accurate methods for the detection and quantification of HAHs, such as polychlorinated dibenzo-p-dioxins (PCDDs), dibenzofurans (PCDFs) and biphenyls (PCBs) and related chemicals in environmental, biological, food and other matrices. This invention provides an improved CALUX cell-based reporting system meeting this need. The current invention outperforms currently available cell bioassay systems in that it can detect dioxins and related dioxin-like chemicals at substantially lower concentrations (10-100 fold lower) and a significantly higher assay response than that of other bioanalytical systems.
Related Materials Additional Technologies by these Inventors Tech ID/UC Case 21033/2009-507-0 Related Cases 2009-507-0
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