The 32P-PostlabeIing Protocols for Assaying Levels of Hydrophobic DNA Adducts in Fish

The Environmental Conservation (EC) Division of the Northwest Fisheries Science Center is evaluating biochemical parameters for use as markers of chemical contaminant exposure and early physiological effects induced by such exposure. A very promising approach is the use of the 32P-postlabeling assay for determining levels of hydrophobic aromatic compounds bound to DNA (DNA adducts) in marine organisms. Recent publications from the EC Division have shown that the 32P-postlabeling method can be used to detect and measure the levels of DNA modified by environmental genotoxic compounds in feral fish. These studies have shown that 1) the levels of hepatic DNA adducts in wild fish positively correlate with the concentrations of polycyclic aromatic hydrocarbons (PAHs) present in marine sediments, and 2) that a strong positive correlation is observed between sediment concentrations of PAHs and the prevalence of neoplastic lesions in liver of marine flatfish. In addition, laboratory studies with model PAHs and sediment extracts have shown that PAH-DNA adducts formed are persistent and have chromatographic characteristics similar to adducts detected in wild fish. These findings suggest that the levels of hepatic DNA adducts found in fish tissues may function as molecular dosimeters of exposure to potentially genotoxic environmental contaminants, such as carcinogenic PAHs. The 32P-postlabeling assay is currently being used as a marker of exposure to potentially genotoxic contaminants in environmental monitoring studies, such as the National Benthic Surveillance Project of NOAA’s National Status and Trends (NS&T) Program and in the Bioeffects Surveys of NOAA’s Coastal Ocean Program. This NOAA technical memorandum describes in detail the 32P-postlabeling method and its application to marine organisms. 1994 NMFS (National Marine Fisheries Service) NWFSC (Northwest Fisheries Science Center) Library Public Domain 1861