Next generation high throughput DNA damage detection platform for genotoxic compound screening

• Main Authors: Sykora, Peter (Author), Witt, Kristine L. (Author), Revanna, Pooja (Author), Smith-Roe, Stephanie L. (Author), Dismukes, Jonathan (Author), Lloyd, Donald G. (Author), Sobol, Robert W. (Author), Engelward, Bevin P (Contributor)
• Other Authors: Massachusetts Institute of Technology. Department of Biological Engineering (Contributor)
• Format: Article
• Language: English
• Published: Nature Publishing Group, 2018-08-28T19:20:04Z.
• Subjects: Article
• Online Access: Get fulltext

Methods for quantifying DNA damage, as well as repair of that damage, in a high-throughput format are lacking. Single cell gel electrophoresis (SCGE; comet assay) is a widely-used method due to its technical simplicity and sensitivity, but the standard comet assay has limitations in reproducibility and throughput. We have advanced the SCGE assay by creating a 96-well hardware platform coupled with dedicated data processing software (CometChip Platform). Based on the original cometchip approach, the CometChip Platform increases capacity ~200 times over the traditional slide-based SCGE protocol, with excellent reproducibility. We tested this platform in several applications, demonstrating a broad range of potential uses including the routine identification of DNA damaging agents, using a 74-compound library provided by the National Toxicology Program. Additionally, we demonstrated how this tool can be used to evaluate human populations by analysis of peripheral blood mononuclear cells to characterize susceptibility to genotoxic exposures, with implications for epidemiological studies. In summary, we demonstrated a high level of reproducibility and quantitative capacity for the CometChip Platform, making it suitable for high-throughput screening to identify and characterize genotoxic agents in large compound libraries, as well as for human epidemiological studies of genetic diversity relating to DNA damage and repair.
National Institutes of Health (U.S.) (Grant CA148629)
National Institutes of Health (U.S.) (Grant ES021116)