A competitive ELISA detecting 7-methylguanosine adduct induced by N-nitrosodimethylamine exposure

Abstract

N-Nitrosodimethylamine is a chemical compound known to be carcinogenic to animals and probably to humans. It is widespread and it can be found in food, tobacco smoke and in industrial emissions, such as in the rubber industry. N-Nitrosodimethylamine exerts its biological effects after metabolic activation by forming methylating nucleic acids in DNA. The most formed adduct is 7-methylguanosine. Our laboratory has developed and validated a competitive enzyme-linked immunosorbent assay in order to detect this adduct in DNA exposed to N-nitrosodimethylamine in vitro or in vivo. The imidazole ring-opening (iro) of 7-methylguanosine was required because of its stability. When 7-methylguanosine iro and serum were incubated at 48C, the assay was 35 times more sensitive than at 378C (50% inhibition at 37 fmol 7-methylguanosine iro per well at 48C and 1.28 pmol at 378C) with a lower limit of detection at 1.58 fmol 7-methylguanosine iro. This assay is reproducible, can be routinely performed and is sensitive enough to detect 7-methylguanosine adduct in DNA samples from human exposed to N-nitrosodimethylamine. We aim to use this method in further studies on epidemiological assessment in people at high risk, such as smokers.