Formalin-fixed paraffin-embedded (FFPE) tissues are rarely used for screening DNA adducts of carcinogens because the harsh conditions required to reverse the formaldehyde-mediated DNA cross-links can destroy DNA adducts. We recently adapted a commercial silica-based column kit used in genomics to manually isolate DNA under mild conditions from FFPE tissues of rodents and humans and successfully measured DNA adducts of several carcinogens including aristolochic acid I (AA-I), 4-aminobiphenyl (4-ABP), and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) (Yun et al. (2013) Anal. Chem.85, 4251–8, and Guo et al. (2016) Anal. Chem.88, 4780–7). The DNA retrieval methodology is robust; however, the procedure is time-consuming and labor intensive, and not amenable to rapid throughput processing. In this study, we have employed the Promega Maxwell 16 MDx system, which is commonly used in large scale genomics studies, for the rapid throughput extraction of DNA. This system streamlines the DNA isolation procedure and increases the sample processing rate by about 8-fold over the manual method (32 samples versus 4 samples processed per hour). High purity DNA is obtained in satisfactory yield for the measurements of DNA adducts by ultra performance liquid chromatography-electrospray-ionization-ion trap-multistage scan mass spectrometry. The measurements show that the levels of DNA adducts of AA-I, 4-ABP, and PhIP in FFPE rodent and human tissues are comparable to those levels measured in DNA from matching tissues isolated by the commercial silica-based column kits and in DNA from fresh frozen tissues isolated by the conventional phenol-chloroform extraction method. The isolation of DNA from tissues is one major bottleneck in the analysis of DNA adducts. This rapid throughput methodology greatly decreases the time required to process DNA and can be employed in large-scale epidemiology studies designed to assess the role of chemical exposures and DNA adducts in cancer risk.

中文翻译：

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从福尔马林固定石蜡包埋的组织中提取DNA的快速通量方法，用于生物监控致癌的DNA加合物。

福尔马林固定石蜡包埋（FFPE）组织很少用于筛选致癌物的DNA加合物，因为逆转甲醛介导的DNA交联所需的苛刻条件会破坏DNA加合物。我们最近对基因组学中使用的基于硅胶的商业化色谱柱试剂盒进行了改造，以在温和条件下从啮齿动物和人类的FFPE组织中手动分离DNA，并成功测量了包括马兜铃酸I（AA-I），4-氨基联苯（4 -ABP），和2-氨基-1-甲基-6-苯基咪唑并[4,5- b ]吡啶（PHIP）（Yun等人（2013）Anal.Chem.31：426-428中。85，4251-8，和Guo等。 （2016）肛门化学88，4780-7）。DNA检索方法可靠。然而，该过程耗时且劳动密集，并且不适合快速通过处理。在这项研究中，我们采用了Promega Maxwell 16 MDx系统，该系统通常用于大规模基因组学研究中，以快速提取DNA。与手动方法相比，该系统简化了DNA分离程序，并使样品处理速率提高了约8倍（32个样品与每小时处理的4个样品）。通过超高效液相色谱-电喷雾-电离-离子阱-多级扫描质谱法测量DNA加合物，可以以令人满意的产率获得高纯度DNA。测量结果表明，AA-1、4-ABP，FFPE啮齿动物和人体组织中的DNA和PhIP与通过商业硅胶色谱柱试剂盒分离的匹配组织的DNA和通过常规苯酚-氯仿提取方法分离的新鲜冷冻组织的DNA中测得的水平相当。从组织中分离DNA是分析DNA加合物的主要瓶颈。这种快速通量的方法极大地减少了处理DNA所需的时间，可用于旨在评估化学暴露和DNA加合物在癌症风险中的作用的大规模流行病学研究。