Detection of carcinogen-DNA adducts by radiommunoassay.

Abstract

COVALENT binding of carcinogen to nucleic acids is believed to be an essential component of the carcinogenic process1, so it is desirable to have highly sensitive and specific methods for detecting such adducts in cells and tissues exposed to known and suspected carcinogens. We describe here a radioimmunoassay (RIA) capable of detecting nanogram amounts of DNA adducts resulting from the covalent binding of the carcinogen N-2-acetylaminofluorene (AAF). AAF and its activated derivative N-acetoxy-AAF (N-Ac-AAF) are potent carcinogens2 and mutagens3,4, and transform cells in culture5,6. DNA obtained from rat liver following in vivo exposure to AAF, and DNA exposed in vitro to N-Ac-AAF contain as the major (80%) adduct N-(deoxyguanosin-8-yl)-acetylaminofluorene (dG-8-AAF)7–9 and a minor (20%) adduct 3-(deoxyguanosin-N2−yl)-acetylaminofluorene (dG-N2-AAF)10. These two types of modification produce markedly different conformational effects on the DNA helix2,11,12. The major adduct, recognised by single strand-specific nucleases in vitro13,14 and DNA repair enzymes in vivo9,15 was used as the immunogen in this study.