YEAST BASED NUCLEAR REPORTER LIFETIME FRET ASSAY SYSTEM

基于酵母的核报告基因寿命 FRET 测定系统

• 批准号: 6151218
• 负责人: Salvador M. Fernandez
• 金额: $ 33.7万
• 依托单位: CIENCIA, INC.
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-08-01 至 2001-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6151218
• 关键词: bioassay; biomedical equipment development; fluorescence resonance energy transfer; method development; scintillation counter; yeasts

We will develop a system consisting of instrumentation, reagents and methods for rapid whole cell nuclear reporter assays for high throughput screening (HTS) of lead compounds in drug discovery applications. The approach is based on the use of green fluorescent proteins as an intracellular market, and fluorescence lifetime sensing to quantify reporter gene expression. A key innovation of the approach is the use of a new fluorescence lifetime sensing technology, recently developed by Ciencia which can be implemented in rugged, low-cost, compact instrumentation, and which is capable of high sensitivity, high lifetime resolution, and rapid measurement time. The proposed lifetime sensing approach will solve some fundamental problems of current intensity-based fluorescence system, such as artifacts due to photobleaching and non- specific cell autofluorescence background, and interference from colored compounds in the sample matrix. the long-term objective is to develop compact, robust, sensitive, low-cost instrumentation and assays for high throughput screening in drug discovery. the proposed work addresses the needs of pharmaceutical companies which are under competitive pressures to bring successful new therapeutic products to market faster and more efficiently. To our knowledge, this would be the first time that fluorescence lifetime sensing is exploited for high throughput screening applications. PROPOSED COMMERCIAL APPLICATION: High throughput screening in lead drug discovery, cell and molecular biology research instrumentation, clinical diagnostics, agriculture, food processing , environmental analysis forensics.

我们将开发一个系统，包括仪器，试剂和 用于高通量的快速全细胞核报告基因测定的方法 在药物发现应用中的先导化合物筛选（HTS）。的 该方法基于使用绿色荧光蛋白作为 细胞内销售和荧光寿命传感来量化 报告基因表达。该方法的一个关键创新是使用 一种新的荧光寿命传感技术，最近开发的 通过Cipher，它可以在坚固，低成本，紧凑的 仪器，并能够高灵敏度，高寿命 分辨率和快速测量时间。所提出的寿命传感 方法将解决当前基于强度的一些基本问题， 荧光系统，如由于光漂白和非荧光 特异性细胞自发荧光背景和有色干扰 样品基质中的化合物。长期目标是发展 紧凑、坚固、灵敏、低成本的仪器和分析， 药物发现中的通量筛选。拟议的工作涉及 制药公司的需求，在竞争压力下， 将成功的新治疗产品更快地推向市场， 有效地据我们所知，这将是第一次 利用荧光寿命传感进行高通量筛选 应用. 拟定商业应用： 先导药物发现、细胞和分子的高通量筛选 生物研究仪器，临床诊断，农业， 食品加工，环境分析和法医学