Overview

The MultiFlow® assay is a mode of action (MOA) assay in TK6 cells to evaluate potential genotoxic response to a test article or its metabolites. By assessing nuclear translocation and binding of p53, the assay discriminates genotoxicants from non-genotoxicants. Furthermore, double strand DNA breaks and mitotic checkpoint delays (G2/M block) results in persistent γH2AX foci and phosphorylation of H3, respectively (further discriminating clastogenicity from aneugenicity).

MultiFlow® Assay Highlights

  • This assay can be used to screen libraries of compounds for potential genotoxic effects (using as little as 5 mg or 200 µL of a 100 mM solution), or to follow up positive mammalian cell assay test results (e.g., micronucleus) when evidence for a non-DNA reactive (e.g., Ames negative) MOA is needed.
  • Up to four compounds can be analyzed concurrently in a series of 20 concentrations, with data acquired by flow cytometry 4- and 24-hours after treatment. Polyploidy, γH2AX shifts, population doubling, H3P+ mitotic cells, and p53 shifts are evaluated in nuclei after cellular lysis.
  • Using three different machine learning algorithms, the probability of genotoxicity, occurring via a clastogenic and/or aneugenic MOA is determined, based upon learning models generated from 24 chemicals spanning multiple MOA and molecular initiating events. For complex results, inter-laboratory cut-off values may be used for each endpoint to assess biological relevance.