The Further Development of a Mammalian DNA Alkaline Unwinding Bioassay With Potential Application to Hazard Identification for Contaminants from Environmental Samples

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Abstract
Recently, we have detailed a DNA alkaline unwinding assay (DA UA) that can be used to rapidly measure chemically induced strand breaks in mammalian cells (Daniel et al., 1985). In this paper we present further development of this assay, including: (1) studies on the relationship between DNA adducts and DNA strand breaks; (2) evaluation of the role of cytotoxicity in DNA strand breaks; and (3) application of the DA UA to cell preparations from the liver of mice dosed with methylating agents. The level of DNA adducts produced in human CCRF-CEM cells by treatment with benzo(a)pyrene diol-epoxide (BPDE), N-acetoxy-2-acetyl amino fluorene (AAAF), and various methylating agents was linear with concentration over several orders of magnitude. Likewise, the level of strand breaks increased with the concentration over the same dose range. The strand breaks/adduct ratio ranged from 0.05 for the methyl adducts to 0.001 for the BPDE adducts. Using these values and the inherent sensitivity of the DA UA (circa 100 to 1000 breaks/ cell), (Daniel et al., 1985), the ability of the assay to detect DNA damage induced by various classes of chemical carcinogens can be calculated.