Objective To improve the chemically-activated luciferase expression (CALUX)bioassay for detection of dioxin-like chemicals (DLCs) based on the toxicity mechanisms ofDLCs. Method A recombinant vector was construc...Objective To improve the chemically-activated luciferase expression (CALUX)bioassay for detection of dioxin-like chemicals (DLCs) based on the toxicity mechanisms ofDLCs. Method A recombinant vector was constructed and used to transfect humanhepatoma (HepG2). The expression of this vector was 10-100 folds higher than that of pGL2used in previous experiments. The transfected cells showed aromatic hydrocarbon receptor(AhR)-meditated luciferase gene expression. The reliability of luciferase induction in thiscell line as a reporter of AhR-mediated toxicity was evaluated, the optimal detection timewas examined and a comparison was made by using the commonly used ethoxyresoufin-O-deethylase (EROD) activity induction assay. Result The results suggested that theluciferase activity in recombinant cells was peaked at about 4 h and then decreased to astable activity by 14 h after TCDD treatment. The detection limit of this cell line was0.11pmol/L, or 10-fold lower than in previous studies, with a linear range from 1 to 100pmol/L, related coefficient of 0.997, and the coefficient of variability (CV) of 15-30%.Conclusion The luciferase induction is 30-fold more sensitive than EROD induction, thedetection time is 68 h shorter and the detection procedure is also simpler.展开更多
文摘Objective To improve the chemically-activated luciferase expression (CALUX)bioassay for detection of dioxin-like chemicals (DLCs) based on the toxicity mechanisms ofDLCs. Method A recombinant vector was constructed and used to transfect humanhepatoma (HepG2). The expression of this vector was 10-100 folds higher than that of pGL2used in previous experiments. The transfected cells showed aromatic hydrocarbon receptor(AhR)-meditated luciferase gene expression. The reliability of luciferase induction in thiscell line as a reporter of AhR-mediated toxicity was evaluated, the optimal detection timewas examined and a comparison was made by using the commonly used ethoxyresoufin-O-deethylase (EROD) activity induction assay. Result The results suggested that theluciferase activity in recombinant cells was peaked at about 4 h and then decreased to astable activity by 14 h after TCDD treatment. The detection limit of this cell line was0.11pmol/L, or 10-fold lower than in previous studies, with a linear range from 1 to 100pmol/L, related coefficient of 0.997, and the coefficient of variability (CV) of 15-30%.Conclusion The luciferase induction is 30-fold more sensitive than EROD induction, thedetection time is 68 h shorter and the detection procedure is also simpler.
