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Many xenobiotics exert their toxic effects through interaction with DNA in the cells of the exposed organism. This interaction may lead to the formation DNA adducts. Some of these may give rise to mutations that initiate cell transformation and, ultimately, the formation of tumors. Sensitive methods for determining DNA adducts are indispensable for the study of chemical mutagenesis and carcinogenesis and for biomonitoring human exposure to genotoxic agents. Alkylating agents form an important class of genotoxic compounds. They react preferentially at the N7-position of guanine. Under neutral or acidic conditions, the adducts can be readily released from the DNA backbone as the free base N7-alkylguanine (N7-AlkGua). The imidazole ring of N7-alkyldeoxyguanosine (N7-AlkdGuo) can be opened under alkaline conditions, which results in formation of a more stable adduct in DNA. To develop immunochemical methods for the detection of N7-alkylations, we immunized mice with various alkylguanosines in the ring-opened form (RON7-AlkdGuo). Antibodies were selected to detect adducts in isolated DNA by competitive ELISA and in single cells by immunofluorescence microscopy (IFM). Various monoclonal antibodies were characterized in detail with respect to specificity and sensitivity toward methylated, ethylated, and hydroxyethylated DNAs. The antibodies showed extensive cross-reactivity toward N7-(m)ethyl- and N7-(2-hydroxyethyl)guanine modifications in the ring-opened form. The limits of detection in the direct and competitive ELISA were 5-10 and 1-2 adducts per 10(6) nucleotides, respectively. The detection limit of the IFM method was about 20 adducts per 10(6) nucleotides(ABSTRACT TRUNCATED AT 250 WORDS)

environmental health perspectives

Determining N7-alkylguanine adducts by immunochemical methods and HPLC with electrochemical detection: applications in animal studies and in monitoring human exposure to alkylating agents.