title: Development of Species-Specific Ah Receptor-Responsive Third Generation CALUX Cell Lines with Enhanced Responsiveness and Improved Detection Limits.
abstract: The Ah receptor (AhR)-responsive CALUX (chemically activated luciferase expression) cell bioassay is commonly used for rapid screening of samples for the presence of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, dioxin), dioxin-like compounds, and AhR agonists/antagonists. By increasing the number of AhR DNA recognition sites (dioxin responsive elements), we previously generated a novel third generation (G3) recombinant AhR-responsive mouse CALUX cell line (H1L7.5c3) with a significantly enhanced response to DLCs compared to existing AhR-CALUX cell bioassays. However, the elevated background luciferase activity of these cells and the absence of comparable G3 cell lines derived from other species have limited their utility for screening purposes. Here, we describe the development and characterization of species-specific G3 recombinant AhR-responsive CALUX cell lines (rat, human, and guinea pig) that exhibit significantly improved limit of detection and dramatically increased TCDD induction response. The low background luciferase activity, low minimal detection limit (0.1 pM TCDD) and enhanced induction response of the rat G3 cell line (H4L7.5c2) over the H1L7.5c3 mouse G3 cells, identifies them as a more optimal cell line for screening purposes. The utility of the new G3 CALUX cell lines were demonstrated by screening sediment extracts and a small chemical compound library for the presence of AhR agonists. The improved limit of detection and increased response of these new G3 CALUX cell lines will facilitate species-specific analysis of DLCs and AhR agonists in samples with low levels of contamination and/or in small sample volumes.
authors.affiliation: Department of Environmental Toxicology, Meyer Hall, University of California , Davis, California 95616, United States.
authors.forename: Jennifer C
authors.initials: JC
authors.lastname: Brennan
authors.affiliation: Department of Environmental Toxicology, Meyer Hall, University of California , Davis, California 95616, United States.
authors.forename: Guochun
authors.initials: G
authors.lastname: He
authors.affiliation: Division of Foods, National Institute of Health Sciences , Kamiyoga 1-18-1, Setagaya-ku, Tokyo 158-8501, Japan.
authors.forename: Tomoaki
authors.initials: T
authors.lastname: Tsutsumi
authors.affiliation: Department of Environmental Toxicology, Meyer Hall, University of California , Davis, California 95616, United States.
authors.forename: Jing
authors.initials: J
authors.lastname: Zhao
authors.affiliation: Center for Coastal Environmental Health and Biomolecular Research, USDOC/NOAA/NOS/NCCOS, Charleston, South Carolina 29412, United States.
authors.forename: Edward
authors.initials: E
authors.lastname: Wirth
authors.affiliation: Center for Coastal Environmental Health and Biomolecular Research, USDOC/NOAA/NOS/NCCOS, Charleston, South Carolina 29412, United States.
authors.forename: Michael H
authors.initials: MH
authors.lastname: Fulton
authors.affiliation: Department of Environmental Toxicology, Meyer Hall, University of California , Davis, California 95616, United States.
authors.forename: Michael S
authors.initials: MS
authors.lastname: Denison
mesh_terms: D000818:Animals; D006528:Carcinoma, Hepatocellular / Q000378:metabolism; D002460:Cell Line; D062126:Databases, Chemical; D019015:Geologic Sediments; D006168:Guinea Pigs; D006801:Humans; D057230:Limit of Detection; D008113:Liver Neoplasms / Q000378:metabolism; D008156:Luciferases / Q000378:metabolism*; D051379:Mice; D010575:Pesticides / Q000032:analysis; D000072317:Polychlorinated Dibenzodioxins / Q000032:analysis; D051381:Rats; D018336:Receptors, Aryl Hydrocarbon / Q000378:metabolism*; D013045:Species Specificity; D014162:Transfection
chemical_list: D010575:Pesticides; D000072317:Polychlorinated Dibenzodioxins; D018336:Receptors, Aryl Hydrocarbon; D008156:Luciferases