CCD-BASED FLUORESCENCE POLARIZATION ASSAY PLATE READER

制药公司目前筛选了数百万种化合物，寻找有前景的候选药物，如与细胞受体具有高亲和力的药物。许多被应用于这一挑战的结合分析(通常是基于荧光的)都存在限制其使用的缺陷。一种检测荧光偏振(FP)变化的技术与其他分析方法相比有几个优点，包括改进了配体和受体之间结合亲和力的量化，以及由于其均匀的(混合和读取)格式而增加了简单性和降低了成本。然而，目前可用的基于FP的仪器并不理想，因为它们的灵敏度低，操作相对较慢。CRI开发的“对称FP”方法将大大提高分析仪器的灵敏度和速度，并将基于FP的分析带入高通量筛查方法的前沿。我们建议继续开发FP高密度平板阅读器。第一阶段和后续工作证明，与现有方法相比，使用标准塑料微量滴定板和低至1 MU1的样品体积，灵敏度提高了100倍，读板速度提高了4倍以上。该仪器将特别有助于开发针对体内低水平表达的具有临床意义的细胞受体的药物。 建议的商业应用： 我们的平板阅读器将用于制药公司的药物开发。规模较大的公司，其中至少有50家，在任何给定的时间都有20到50个正在进行的药物筛选计划。我们预计，他们现有的大多数配体筛选仪器将被更高吞吐量、更具成本效益的仪器所取代，特别是拟议的CRI FP平台。保守估计，潜在市场价值为600-1800万美元。

Pharmaceutical companies currently screen millions of compounds looking for promising drug candidates such as agents that bind with high affinity to cellular receptors. Many binding assays (often fluorescence-based) being applied to this challenge suffer from drawbacks that limit their use. A technique examining changes in fluorescence polarization (FP) promises several advantages over other assays, including improved quantification of binding affinity between ligand and receptor, and increased simplicity and lower cost, due to its homogeneous (mix-and-read) format. Currently available FP-based instruments are not ideal, however, because of their low sensitivity and relatively slow operation. An approach, "Symmetric FP", developed at CRI, will lead to analytical instruments with greatly improved sensitivity and speed and should bring FP-based assays to the forefront of high-throughput screening methods. We propose to continue development of an FP high-density plate reader. Phase I and subsequent work have demonstrated 100-fold improvement in sensitivity using standard plastic microtitre plates and sample volumes as low as 1 mu1, and a more than 4-fold increase in plate-reading speed, compared to existing methods. The instrument will be particularly useful for developing drugs targeting clinically significant cellular receptors expressed at low levels in vivo. PROPOSED COMMERCIAL APPLICATION: Our plate reader would be used for drug discovery by pharmaceutical companies. The larger companies, of which there are at least 50, have between 20 and 50 drug screening programs ongoing at any given time. We expect that most of their existing ligand-screening instrumentation will be replaced by higher throughput, more cost-effective instruments, notably the proposed CRI FP platform. Conservative estimates suggest a potential market for 6-18 million dollars.