PFI-TT: Developing an integrated platform for high accuracy measurements of viral particle count and infectious titer

PFI-TT：开发用于高精度测量病毒颗粒计数和感染滴度的集成平台

• 批准号: 2141135
• 负责人: Sara Rouhanifard
• 金额: $ 24.98万
• 依托单位: Northeastern University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-02-01 至 2024-01-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2141135&HistoricalAwards=false
• 关键词: PFI; TT; Developing; integrated; platform

The broader impact/commercial potential of this Partnerships for Innovation - Technology Translation (PFI-TT) project is to measure viral concentrations in clinical or research settings. Select vaccines and gene therapies are delivered via viral vectors, but the precise dosing is poorly known, leading to quality control concerns. Current measurement techniques are indirect, imprecise, and nonstandard; and they require time and specialized equipment. Accurate measurement of viral particles is critical for precise benchmarking of diagnostics and appropriate dosing of therapeutics. This project develops a new technology to measure viral concentrations and infectivity. The proposed project addresses viral quantification techniques as they suffer from several limitations. Typical infectivity assays are highly dependent on the specific cell line used for viral infection, leading to high variability between labs. Furthermore, natural viruses are not engineered to contain a fluorescent transgene for easy identification, and some particles will not contain the viral genome - leading to competition between those containing the gene “payload” and empty vectors. In general, the density, infectivity, and replication efficiency of viral particles in a given sample is poorly known. This project advances a technology with three functioins: quantifying the total number of particles, measuring genome-containing units, and assessing infectivity of a given sample of the virus. The new method combines nano-pore technology using biophysical properties to identify viral particles with single-molecule RNA fluorescence in situ hybridization to discriminate down to single molecules of viral RNA in infected cells.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

创新-技术转化伙伴关系（PFI-TT）项目的更广泛影响/商业潜力是测量临床或研究环境中的病毒浓度。某些疫苗和基因疗法是通过病毒载体提供的，但精确的剂量鲜为人知，导致质量控制方面的担忧。目前的测量技术是间接的、不精确的和不标准的；而且他们需要时间和专门的设备。病毒颗粒的精确测量对于精确的诊断基准和适当的治疗剂量至关重要。该项目开发了一种测量病毒浓度和传染性的新技术。拟议的项目解决了病毒量化技术，因为它们受到几个限制。典型的传染性测定高度依赖于用于病毒感染的特定细胞系，导致实验室之间的高度可变性。此外，天然病毒没有被设计成含有易于识别的荧光转基因，而且一些颗粒将不包含病毒基因组——导致含有基因“有效载荷”和空载体之间的竞争。一般来说，在一个给定的样本中，病毒颗粒的密度、传染性和复制效率是鲜为人知的。该项目推进了一项具有三种功能的技术：量化颗粒总数，测量包含基因组的单位，以及评估给定病毒样本的传染性。新方法结合纳米孔技术，利用生物物理特性识别病毒颗粒，并结合单分子RNA荧光原位杂交技术，在感染细胞中区分到病毒RNA的单分子。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。