The ToxTracker assay is a state-of-the-art stem cell-based reporter assay that provides mechanistic insight into genotoxic properties of compounds. For screening purposes, a selection of the ToxTracker biomakers is applied to test a large number of compounds for genotoxicity. Activation of the Bscl2-GFP biomarker for DNA damage has a very high correlation with the Ames test. The Rtkn-GFP biomarker is activated by DNA strand breaks and strongly correlates with the regulatory clastogenicity assays (MN/CA). Combining these genotoxic markers will result in an assay which combines mutagenic and clastogenic endpoints in a fast format and requires little compound.
Key Features
- screen up to hundreds of compounds per week
- excelent predictor for genotoxicity
- small amount of compound required (0,25-0,5 mg)
- quick turn around – report in a week
- metabolic activation using S9 liver extract
- combines clastogenic and mutagenic endpoints
Combined biomarkers for Genotoxicity
ToxTracker® is a mammalian stem cell-based reporter assay that detects activation of specific cellular signalling pathways upon chemical exposure. ToxTracker contains six different GFP-tagged reporter cell lines that together allow the discrimination between induction of DNA damage, oxidative stress and/or protein damage in a single test. Genotoxicity is detected by the Bscl2-GFP reporter for pro-mutagenic DNA lesions and DNA replication stress, and the Rtkn-GFP reporter for DNA double strand breaks.
For screening purposes, a selection of biomakers can be applied to test a large number of compounds for genotoxicity. Activation of the Bscl2-GFP biomarker for DNA damage has a very high correlation with the Ames test (figure 1). The Rtkn-GFP biomarker is activated by DNA strand breaks and strongly correlates with the regulatory clastogenicity assays (MN/CA) (figure 2). Combining these genotoxic markers will result in an assay which combines mutagenic and clastogenic endpoints in a fast format and requires little compound.
Figure 1: Comparison between activation of the Bscl2-GFP genotoxicity reporter activation and the Ames and/or MLA mutation assays for the ECVAM library of reference compounds.
Figure 2: Comparison between activation of the Rtkn-GFP genotoxicity reporter activation and the in vitro and in vivo micronucleus and/or chromosome aberration tests for the ECVAM library of reference compounds.
Of course we can adjust the selection of biomarker genes we analyse, for example to include or focus on oxidative stress. Please contact us to discuss your needs.
Screening set-up
- Screening in 384-wells plates
- Compounds provided by customer as stock solution
- Start with dose-range finding (7 concentrations, 10-fold dilution steps)
- Miniturized ToxTracker (5 concentrations, 2 fold dilution steps)
- Automated data analysis
Reporting
A full report will be created at the end of the project. All test results are summarised in an excel spreadsheet. Data is summarised in a color heat map for rapid identification of positive test results.
Example heatmap:
