Improvement of chemically-activated luciferase gene expression bioassay for detection of dioxin-like chemicals / 生物医学与环境科学(英文)
Biomedical and Environmental Sciences
; (12): 58-66, 2002.
Article
em En
| WPRIM
| ID: wpr-264332
Biblioteca responsável:
WPRO
ABSTRACT
<p><b>OBJECTIVE</b>To improve the chemically-activated luciferase expression (CALUX) bioassay for detection of dioxin-like chemicals (DLCs) based on the toxicity mechanisms of DLCs.</p><p><b>METHODS</b>A recombinant vector was constructed and used to transfect human hepatoma (HepG2). The expression of this vector was 10-100 folds higher than that of pGL2 used in previous experiments. The transfected cells showed aromatic hydrocarbon receptor (AhR)-meditated luciferase gene expression. The reliability of luciferase induction in this cell line as a reporter of AhR-mediated toxicity was evaluated, the optimal detection time was examined and a comparison was made by using the commonly used ethoxyresoufin-O-deethylase (EROD) activity induction assay.</p><p><b>RESULTS</b>The results suggested that the luciferase activity in recombinant cells was peaked at about 4 h and then decreased to a stable activity by 14 h after TCDD treatment. The detection limit of this cell line was 0.11 pmol/L, or 10-fold lower than in previous studies, with a linear range from 1 to 100 pmol/L, related coefficient of 0.997, and the coefficient of variability (CV) of 15-30%.</p><p><b>CONCLUSION</b>The luciferase induction is 30-fold more sensitive than EROD induction, the detection time is 68 h shorter and the detection procedure is also simpler.</p>
Texto completo:
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Base de dados:
WPRIM
Assunto principal:
Patologia
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Farmacologia
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Bioensaio
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Células Tumorais Cultivadas
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Transfecção
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Indução Enzimática
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Regulação da Expressão Gênica
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Sensibilidade e Especificidade
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Carcinoma Hepatocelular
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Citocromo P-450 CYP1A1
Tipo de estudo:
Diagnostic_studies
Limite:
Humans
Idioma:
En
Revista:
Biomedical and Environmental Sciences
Ano de publicação:
2002
Tipo de documento:
Article