Comet assay in murine bone-marrow cell line (FDC-P2)

Toxicology in Vitro : an International Journal Published in Association with BIBRA
K OshidaY Miyamoto

Abstract

The comet assay, also known as the single cell gel electrophoresis (SCGE) assay, is a rapid, simple, visual and sensitive technique for measuring DNA damage in mammalian cells. In the present study, Methyl methanesulfonate (MMS), 4-Nitrosoquinoline-Oxide (4NQO), Cyclophosphamide (CPA), and Benzo(a)pyrene (BP)-induced DNA damage was assayed in vitro in a murine bone-marrow cell line (FDC-P2), with or without an activation mixture (rat liver S9). All compounds caused significant DNA damage. With MMS and 4NQO, the frequency of comet tails, scored manually under a fluorescence microscope, increased dose-dependently, and reached a maximum of 53.2 and 74.8% respectively. Three parameters indicating DNA damage in the comet assay with the two-layer method, tail length, %DNA in tail, and tail moment, calculated using the automated image analysis software "Comet Analyzer v1.5" increased with all compounds. With MMS and 4NQO, all parameters increased at concentrations over 40 and 0.25 micromol/L, respectively. The in vitro comet assay with rat liver S9 could detect DNA damage caused by the metabolites of CPA and BP. The comet assay using the two-layer method is easy and efficient, and so can be conducted on a routine as basis. The assay w...Continue Reading

References

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