FORMATION AND REMOVAL OF SPECIFIC ACETYLAMINOFLUORENE DNA ADDUCTS IN MOUSE AND HUMAN-CELLS MEASURED BY RADIOIMMUNOASSAY

Abstract

Rabbit antiserum prepared against N-(guanosin-8-yl)acetylaminofluorene was utilized in a radioimmunoassay to detect formation and removal of C-8 adducts from the DNA of cultured cells exposed to N-acetoxy-2-acetylaminofluorene. The assay was able to quantitate both acetylated and deacetylated C-8 adducts between 0.5 and 5 pmol while the N2 adduct, 3-(deoxyguanosin-N2-yl)acetylaminofluorene, was not detected below 160 pmol. By varying the proportions of acetylated and deacetylated C-8 adducts in the radioimmunoassay, a series of standard curves were developed from which the relative proportion of each adduct could be determined in unknown mixtures. DNA from mouse epidermal cells and human skin fibroblasts exposed to N-acetoxy-2-acetylaminofluorene in culture contained only 3 and 5%, respectively, of the C-8 adduct in the acetylated form. Quantitation by radioimmunoassay of total C-8 adducts bound to DNA yielded values .apprx. 25% lower than total carcinogen binding determined by radiolabeling. When removal of C-8 adducts was followed over a 23 h, carcinogen-free culture period, mouse and human cells removed 40 and 50%, respectively, of bound acetylated and deacetylated C-8 adducts. Radioimmunoassay is a versatile molecular probe for studies of chemical carcinogens.