Flow cytometry-based Unscheduled DNA Synthesis (FLUDS)

描述(申请人提供)：工业上越来越需要一种廉价的筛选分析方法，能够可靠地确定每年合成的大量新化合物的遗传毒性潜力。非程序DNA合成(UDS)分析是一种鉴定和表征遗传毒性化学物质的成熟方法。然而，标准化验的几个技术方面限制了其广泛使用。在第一阶段研究中，我们通过加入流式细胞术作为检测方法，对标准的UDS分析进行了修改，并证明了我们的优化版本UDS，称为FLUDS的可行性。FLUDS检测允许更高的吞吐量和更短的研究时间(标准UDS只需几天而不是几周)。这种新的检测方法可以区分天然的和生物转化的基因毒物，而且是完全定量的。它可靠地预测了乙基亚硝脲、除草剂、2-乙酰氨基荧烯和荧烯引起的UDS。我们打算通过测试一组扩大的遗传毒性和非遗传毒性化学物质来预先验证FLUDS检测，为ICCVAM指南下的多实验室验证做准备。对于制药、化工、化妆品和消费品行业的客户来说，预先验证的FLUDS检测将具有相当大的商业价值。

DESCRIPTION (provided by applicant): There is a growing need in industry for an inexpensive screening assay which can reliably determine the genotoxic potential of the large number of new chemical compounds synthesized each year. The Unscheduled DNA Synthesis (UDS) Assay is a proven method to identify and characterize genotoxic chemicals. However, several technical aspects of the standard assay limit its widespread use. We have made modifications to the standard UDS assay in Phase I studies through incorporation of flow cytometry as a detection method, and have proven feasibility of our optimized version of UDS, termed FLUDS. The FLUDS assay allows higher throughput and much shorter study times (a few days versus weeks for the standard UDS). This new assay can discriminate between innate and biotransformed genotoxicants, and is fully quantitative. It has reliably predicted the UDS induced by ethylnitrosourea, aphidicolin, 2- acetylaminofluorene, and fluorene. We intend to pre-validate the FLUDS assay by testing an expanded set of genotoxic and non-genotoxic chemicals in preparation for a multi-laboratory validation under ICCVAM guidelines. The pre-validated FLUDS assay would be of considerable commercial value to customers in the pharmaceutical, chemical, cosmetic and consumer products industries.