Title: Application of the CALUX (chemically activated luciferase gene expression) assay for measuring TCDD-equivalents in sediment, pore water, and blood samples

Auteur(s): Murk AJ ; Jonas A ; Brouwer A ; Leonards PEG ; Denison MS ;  ; 

Year: 1996

Journal: Organohalogen Compounds

Subject(s): CALUX ; gene expression ; Luciferase ; bioassay ; Ah receptor ; sediment ; blood plasma ; 2,3,7,8-tetrachlorodibenzo-p-dioxin ; TCDD ; dioxin-like compounds ; extraction ; clean-up ; PCB ; EROD ;  ; 

Summary: The results presented in this paper demonstrate that the in vitro CALUX (chemical activated luciferase gene expression) bioassay is a rapid sensitive assay for assessing the toxic potency of (mixtures of) Ah-receptor active compounds in sediment, pore water and blood plasma. Recombinant rat (H4IIE) and mouse (Hepalclc7) hepatoma cell lines, containing arylhydrocarbon receptor (AhR)-mediated luciferase gene expression, were used to determine the CALUX-response of sample extracts. Using a 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) standard curve, the CALUX responses for tbe samples were converted into so-called CALUX-TEQs (TCDD-equivalents). The detection limit of the CALUX assay was 0.5 fmol of TCDD-equivalents. The CALUX activity induced by organic extracts from 450 mg aliquots of sediment or 250 µl aliquots of pore-water corresponded well with the instrumentally analysed level of dioxin-like compounds in the sediment. Using pore water, only a simple and rapid extraction procedure was needed, without additional clean-up to prevent cell death. The CALUX response both for the fatty, uncleaned organic extract and the cleaned extract of blood plasma from experimentally exposed eider ducks, correlated significantly with the PCB levels measured in abdominal fat or blood plasma, and with the hepatic EROD activity in the same animals. An important advantage of the CALUX-assay compared to the ethoxyresorufin O-deethylase (EROD) assay is that the CALUX assay is insensitive to substrate inhibition, which is demonstrated in this paper for 3.3',4,4',5-pentachlorobiphenyl (PCB-126).