Application of the Lux-Fluoro test as bioassay for combined genotoxicity and cytotoxicity measurements by means of recombinant Salmonella typhimurium TA1535 cells

The Lux-Fluoro test is a bioassay, which coincidentally measures the cytotoxic and genotoxic potency of a given substance based on the receptor reporter principle. For genotoxicity testing, bioluminescence occurs as a response to the presence of DNA-damaging agents. It is brought about by the induction of the promoterless luxCDABFE genes of Photobacterium leiognathi as reporter component under the control of a strong SOS promoter as receptor component. At concentrations of DNA-damaging agents which only scarcely affect cell survival, a high level of light production is induced. For testing cytotoxic agents viability of the bacteria is included as a test parameter describing the cytotoxicity of the agents. For that reason, Salmonella typhimurium TA1535 cells transformed with the bacterial protein expression vector pGFPuv were employed. This plasmid controls green fluorescent protein (GFP) expression by the lac promoter in TA1535 cells constitutively. A panel of recombinant S. typhimurium strains carrying either the SOS-Lux-plasmid (TA1535-pPLS-1) or the fluorescence mediating lac-GFPuv plasmid (TA1535-pGFPuv) was used to record in parallel agents (mitomycin C, chloramphenicol, doxorubicin hydrochloride, bleomycin sulphate and hydrogen peroxide) that effect the genetic material conveying either genotoxic, cytotoxic or geno- and cytotoxic effects. The light and fluorescence transmissions of untreated and chemical-treated cells were measured in a microtiter plate reader and luminescence induction factors (F-i) as well as fluorescence deduction factors (Fd) were calculated for the genotoxic and cytotoxic potential of the applied agents. (C) 2001 Elsevier Science B.V. All rights reserved.