High Sensitivity Detection of Viral DNAs in Cancer Cells

描述（由申请人提供）：几种人类癌症，包括宫颈癌 癌症是由病毒感染引起的。致癌的分子机制 通常涉及病毒介导的正常细胞周期和生长的丧失 由于病毒整合到宿主基因组中而引起的调节。在某些情况下， 整合可以仅涉及病毒DNA的几个拷贝到单个拷贝。在 原位聚合酶链反应（IS-PCR）提供了高灵敏度的检测 低拷贝数的病毒DNA，但该测定是繁琐的，并且遭受 再现性和人为问题。酪胺信号 扩增）为基础的原位杂交提供了高灵敏度，但是，像 IS-PCR，样品通量低。第一阶段SBIR提案旨在制定 用于检测低拷贝数病毒DNA的灵敏、高通量方法 通过结合单细胞凝胶微滴（GMD）包封的癌细胞 技术，原位杂交，TSA和流式细胞术。在该方法中，TSA 将用于将杂交信号放大到可通过流动检测的水平 细胞仪除了高检测灵敏度和样品通量之外， 该方法将通过减少样品损失而允许检测罕见的癌细胞 通过GMD封装I期研究将使用宫颈癌作为 实验模型 拟议商业应用：不可用

DESCRIPTION (provided by applicant): Several human cancers, including cervical cancer, are caused by viral infection. Molecular mechanisms of carcinogenesis generally involve a virally mediated loss of normal cell cycle and growth regulation due to viral integration into the host genome. In cases, viral integration can involve only a few copies to a single copy of viral DNA. In situ polymerase chain reaction (IS-PCR) offers high sensitivity for detecting low copy number viral DNA, but the assay is cumbersome and suffers from reproducibility and artifactual problems. TSA (tyramide signal amplification)-based in situ hybridization offers high sensitivity, but, like IS-PCR, sample throughput is low. This Phase I SBIR proposal aims to develop a sensitive, high throughput method for detecting low copy number viral DNA in cancer cells by combining single cell gel microdrop (GMD) encapsulation technology, in situ hybridization, TSA and flow cytometry. In this method, TSA will be used to amplify the hybridization signals to levels detectable by flow cytometry. In addition to high detection sensitivity and sample throughput, this method will permit detection of rare cancer cells by reducing sample loss through GMD encapsulation. Phase I-research will use cervical cancer as an experimental model. PROPOSED COMMERCIAL APPLICATION: NOT AVAILABLE