AFLATOXIN DNA ADDUCTS DETECTED BY MONOCLONAL ANTIBODIES

黄曲霉毒素B1（AFB 1）是已知的最强的肝脏致癌物之一， 许多流行病学研究发现， AFB 1暴露与人类肝癌之间存在强正相关。 这些数据激发了我们开发技术来监测 AFB 1的个体暴露和代谢。 为了实现这一目标， 使用高度特异性的单克隆抗体， 分析和定量黄曲霉毒素代谢物的分析化学方法 在生物样本中。 在以下研究中，我们建议：（1）生产 并表征单克隆抗体以检测主要的黄曲霉毒素-DNA 加合物：AFB 1-N7-Gua和AF-FaPyr。 抗原生产的两种策略 将用于 (A)一种新的DNA加合物-肟抗原和（B）一种制备的半抗原 由合成黄曲霉毒素环氧化物产生的黄曲霉毒素鸟苷加合物 将尝试诱导抗体对DNA的特异性和亲和力 通过先前的半抗原化程序未获得的加合物;（2）开发 分析免疫测定和制备亲和层析方法 足以检测细胞、组织和排泄物中的AFB 1及其衍生物 长期服用黄曲霉毒素B1的动物， 肿瘤发生 由于主要的AFB-N7-Gua的脱嘌呤不稳定性， 排除了使用32 P-后标记方法，高亲和力DNA加合物 必须产生特异性抗体以检测低水平的 非放射性标记的细胞AFB-DNA病变;和（3）评价 （3）中开发的相关剂量测定程序的适用性 对细胞内AFB 1-DNA加合物和代谢物浓度的反应， 组织和排泄物的大鼠单剂量和慢性与黄曲霉毒素B1。 这些 研究将改进方法，以探索饮食之间的对应关系， 摄入黄曲霉毒素与生物标志物，如排泄的AFB-DNA 加合物，以确定这些标记物是否具有必要的性质 对人体暴露状况进行非侵入性评估。

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.