IMAGING ONCOLYTIC ADENOVIRUSES SPREAD IN A 3D SYSTEM

对 3D 系统中传播的溶瘤腺病毒进行成像

• 批准号: 8365770
• 负责人: HUNG Y. FAN
• 金额: $ 0.55万
• 依托单位: UNIVERSITY OF CALIFORNIA-IRVINE
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-08-20 至 2012-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8365770
• 关键词: Adenoviruses; Appearance; Biological Models; Biology; Cells; Data; Fluorescence; Funding; Grant; Image; Individual; Infection; Laboratories; Microscope; Monitor; National Center for Research Resources; Oncolytic; Optics; Principal Investigator; Research; Research Infrastructure; Resource Sharing; Resources; Solid; Source; Suspension substance; Suspensions; System; United States National Institutes of Health; Viral; Virus; cost; matrigel; monolayer; reconstitution; research study; three dimensional structure

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Study of viral spread in 3D cultures. The most relevant system for modeling viral spread would be in solid cell masses. As shown in the preliminary data, we have been able to image individual infected cells in solid masses by reconstituting infected and uninfected cells into spheres on Matrigel. We will first carry out analysis on AdEGFPuci-infected 293 cells. We will first infect a monolayer of 293 cells with AdEGFPuci, and 6 hr after infection, the infected cells will be trypsinized. The infected single cell suspensions will be mixed with an excess of uninfected 293 cells, and plated onto Matrigel matrices to allow formation of cell aggregates and spheres. The spheres will be monitored daily for appearance of green fluorescent cells and fluorescent Z-stack montages will be recorded and compiled for any sphere that initially show one infected cell. A confocal microscope suitable for these experiments is available in the campus optical biology shared resource. It seems possible that the rate of virus spread may be more rapid in 3D structures than in monolayer, or that the number of secondarily infected cells may be larger. It will also be possible to determine the effects of initial MOI on spread through 3D cultures.

该子项目是利用资源的众多研究子项目之一 由 NIH/NCRR 资助的中心拨款提供。子项目的主要支持 并且子项目的首席研究员可能是由其他来源提供的， 包括其他 NIH 来源。 子项目可能列出的总成本 代表子项目使用的中心基础设施的估计数量， NCRR 赠款不直接向子项目或子项目工作人员提供资金。 3D 培养物中病毒传播的研究。模拟病毒传播最相关的系统是固体细胞团。如初步数据所示，我们已经能够通过将感染和未感染的细胞重建为基质胶上的球体，对固体块中的单个感染细胞进行成像。 我们首先对AdEGFPuci感染的293细胞进行分析。我们首先用 AdEGFPuci 感染单层 293 细胞，感染后 6 小时，对感染的细胞进行胰蛋白酶消化。感染的单细胞悬浮液将与过量的未感染的293细胞混合，并铺板到Matrigel基质上以形成细胞聚集体和球体。将每天监测球体是否出现绿色荧光细胞，并将记录并编译最初显示受感染细胞的任何球体的荧光 Z 堆栈蒙太奇。校园光学生物学共享资源中提供了适合这些实验的共焦显微镜。病毒在 3D 结构中的传播速度似乎可能比在单层结构中更快，或者二次感染细胞的数量可能更多。还可以确定初始 MOI 对 3D 培养传播的影响。