AFLATOXIN DNA ADDUCTS DETECTED BY MONOCLONAL ANTIBODIES

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Aflatoxin B1 (AFB1) is one of the most potent liver carcinogens known for experimental animals, and a number of epidemiological studies have found strong positive associations between AFB1 exposure and human liver cancer. These data have spurred our interest to develop technologies to monitor individual exposure to and metabolism of AFB1. To accomplish this goal, highly specific monoclonal antibodies are used with complementary analytical chemistry methods to analyze and quantify aflatoxin metabolites in biological samples. In the following study, we propose to: (1) Produce and characterize monoclonal antibodies to detect the major aflatoxin-DNA adducts: AFB1-N7-Gua and AF-FaPyr. Two strategies for antigen production will be used. (A) A novel DNA adduct-oxime antigen and (B) a hapten made from aflatoxin guanosine adduct produced by synthetic aflatoxin epoxide will be tried to induce an antibody specificity and affinity for the DNA adducts not attained by previous haptenization procedures; (2) Develop analytical immunoassays and preparative affinity chromatographic methods adequate to detect AFB1 and its derivatives in cells, tissues and excreta of animals chronically dosed with aflatoxin B1 at levels which induce tumorigenesis. Since the lability to depurination of the major AFB-N7-Gua precludes the use of 32P-post-labeling methods, high affinity DNA adduct specific antibodies must be produced for the detection of low levels of non-radiolabeled cellular AFB-DNA lesions; and (3) Evaluate the applicability of assay procedures developed in (3) for relating dose response to concentrations of AFB1-DNA adducts and metabolites in cells, tissues and excreta of rats dosed singly and chronically with AFB1. These studies will refine methods to probe the correspondence between dietary intake of aflatoxins with biological markers, such as excreted AFB-DNA adducts, to determine whether these markers have the requisite properties for non-invasive assessment of the exposure status of people.