Vitotox, 10 assays - 3300 Euro/kit

● The Vitotox is a new genotoxicity and cytotoxicity test based on genetically modified test organisms for faster and more superior results. The Vitotox assay, an SOS-bioluminescence Salmonella typhimurium test, uses the light emission of bacteria to detect the genotoxicity, toxicity and mutagenic potency of the sample. Increased luciferase activity is measured kinetically. Effects of cytotoxicity can be measured simultaneously with genotoxicity. The test detects any DNA damage and gene mutations.

Description

● The GENTAUR VitotoxTM 10 Kit is based on the original VitotoxTM test established at the Flemish Institute for Technological Research (van der Lelie et al., 1997, Verschaeve et al. 1999) as a reaction to the conventional bacterial short-term tests to detect genotoxic and toxic agents. The Salmonella typhimurium bacteria used in the VitotoxTM 10 Kit lack the necessary oxidative enzymes for metabolizing foreign compounds to electrophilic metabolites capable of reacting with DNA, therefore the bacteria are treated with the test agent in the presence and absence of a post-mitochondrial supernatant (S9). This supernatant is prepared from the livers of aroclor-treated rats. After being incubated overnight in rich growth medium, a dilution of the bacterial suspension is incubated for one more hour on an orbital shaker in poor growth medium the following day. Coincidently, dilutions of different concentrations of the test agent are made and are transferred to a 96 multiwell plate. Next to the dilution series, a blank of the solvent and a positive control are added (e.g. Bentso(α)pyrene in with S9 and 4-Nitroquinoline-oxide without S9). The genotoxicity and toxicity measurements are performed using a microplate luminometer that enables measurements of emitted light every 5 minutes over a period a 4 hours. After completion the data are transferred to a spreadsheet application and the signal-to-noise ratio (S/N), i.e. the light production of exposed bacteria divided by the light production of non-exposed bacteria, is calculated for each measurement. The S/N is calculated of the Genox and Cytox strain separately, as well as the ratio between the maximum S/N values of the Genox and Cytox strains. Based on these experimental grounds an agent is considered genotoxic, cytotoxic or not.

     ● Very small amount of test compound required

     ● Short assay time (An answer on genotoxicity is obtained in less than 4 hours)

     ● High reproducibility and sensitivity

     ● Good correlation with the government standard Ames Test

     ● Very simple and cost-efficient to perform

     ● Automatic reading of kinetic data

Principle

Two genetically engineered Salmonella typhimurium strains are used in this test system, TA104 recN2-4 (Genox strain) and TA104 pr1 (Cytox strain):

The Genox strain carries a plasmid containing the bacterial luciferase operon (luxCDABE) of luminous bacteria Vibrio fischeri under transcriptional control of a mutated recN promoter (recN2-4) that is part of the SOS-system. Genotoxic compounds activate the recN promoter in the Genox strain, which results in transcriptional induction of the lux operon followed by the enhancement of light emission.

The Cytox strain possesses a plasmid containing the Vibrio fischeri lux operon, transcriptionally controlled by the strong pr1 promoter and thus constitutively produces light. This strain is used as an internal control system as some compounds appear to have a direct influence on the light production or enhance the metabolism of the bacteria creating false-positive results.

The cytotoxicity of the compounds is simultaneously assayed with reference to the Cytox strain to identify the non-specific enhancement of light emission. Concomitant use of the Genox and the Cytox strains allows us to identify false positive results caused by non-specific light emission induced by other mechanisms, and not by the genotoxic effect.

Kit Content

     ● Genox strain S. typhimurium TA104 recN2-4, Five Blue vials of 100 µl - Store at        -70°C

REFERENCES

§ Van der Lelie D, Regniers L, Borremans B, Provoost A, Verschaeve L., The VITOTOX test, an SOS bioluminescence Salmonella typhimurium test to measure genotoxicity kinetics, Mutat Res., 1997 Mar 17;389(2-3):279-90.

§ Verschaeve L, Van Gompel J, Thilemans L, Regniers L, Vanparys P, van der Lelie D., VITOTOX bacterial genotoxicity and toxicity test for the rapid screening of chemicals, Environ Mol Mutagen., 1999;33(3):240-8.