Enhancing the response of CALUX and CAFLUX cell bioassays for quantitative detection of dioxin-like compounds  

作　　者：BASTON David S. KHAN Elaine SORRENTINO Claudio DENISON Michael S. 

机构地区：[1]Laboratorium voor Analytische en Milieu Chemie,Vrije Universiteit Brussel [2]Pesticide Epidemiology Section,Pesticide and Environmental Toxicology Branch,Office of Environmental Health Hazard Assessment [3]Human and Ecological Risk Office,Brownfields and Environmental Restoration Program,Department of Toxic Substances Control [4]Department of Environmental Toxicology,University of California

出　　处：《Science China Chemistry》2010年第5期1010-1016,共7页中国科学（化学英文版）

基　　金：support from the National Institutes of Environmental Health Sciences Superfund Basic Research Grant(ES004699(MSD));the California Agricultural Experiment Station,the Startup Fund of 100 Talents Program of Chinese Academy of Sciences;the National Natural Science Foundation of China (20921063);the National Basic Research Program(2010CB933500)

摘　　要：Reporter genes produce a protein product in transfected cells that can be easily measured in intact or lysed cells and they have been extensively used in numerous basic and applied research applications.Over the past 10 years,reporter gene assays have been widely accepted and used for analysis of 2,3,7,8-tetrachlorodibenzo-p-dioxin and related dioxin-like compounds in various types of matrices,such as biological,environmental,food and feed samples,given that high-resolution instrumental analysis techniques are impractical for large-scale screening analysis.The most sensitive cell-based reporter gene bioassay systems developed are the mechanism-based CALUX(Chemically Activated Luciferase Expression) and CAFLUX(Chemically Activated Fluorescent Expression) bioassays,which utilize recombinant cell lines containing stably transfected dioxin(AhR) responsive firefly luciferase or enhanced green fluorescent protein(EGFP) reporter genes,respectively.While the current CALUX and CAFLUX bioassays are very sensitive,increasing their lower limit of sensitivity,magnitude of response and dynamic range for chemical detection would significantly increase their utility,particularly for those samples that contain low levels of dioxin-like HAHs(i.e.,serum) .In this study,we report that the addition of modulators of cell signaling pathways or modification of cell culture conditions results in significant improvement in the magnitude and overall responsiveness of the existing CALUX and CAFLUX cell bioassays.Reporter genes produce a protein product in transfected cells that can be easily measured in intact or lysed cells and they have been extensively used in numerous basic and applied research applications.Over the past 10 years,reporter gene assays have been widely accepted and used for analysis of 2,3,7,8-tetrachlorodibenzo-p-dioxin and related dioxin-like compounds in various types of matrices,such as biological,environmental,food and feed samples,given that high-resolution instrumental analysis techniques are impractical for large-scale screening analysis.The most sensitive cell-based reporter gene bioassay systems developed are the mechanism-based CALUX(Chemically Activated Luciferase Expression) and CAFLUX(Chemically Activated Fluorescent Expression) bioassays,which utilize recombinant cell lines containing stably transfected dioxin(AhR) responsive firefly luciferase or enhanced green fluorescent protein(EGFP) reporter genes,respectively.While the current CALUX and CAFLUX bioassays are very sensitive,increasing their lower limit of sensitivity,magnitude of response and dynamic range for chemical detection would significantly increase their utility,particularly for those samples that contain low levels of dioxin-like HAHs(i.e.,serum) .In this study,we report that the addition of modulators of cell signaling pathways or modification of cell culture conditions results in significant improvement in the magnitude and overall responsiveness of the existing CALUX and CAFLUX cell bioassays.

关 键 词：DIOXIN REPORTER gene BIOASSAY CALUX CAFLUX 

分 类 号：X830[环境科学与工程—环境工程]