Microarraying and Screening Chemical Libraries

描述(由申请人提供)：反应生物学公司(宾夕法尼亚州马尔文)已经开发出均相反应微阵列，以服务于NIH路线图倡议，在药物发现、大规模IC50测定和选择性分析中进行纳升规模的生化高通量筛选(HTS)。这些纳升反应比目前使用的井板格式小1000到10000倍。提供了针对丝氨酸蛋白酶、半胱氨酸氨基转移酶、基质金属蛋白酶、酪氨酸激酶和丝氨酸/苏氨酸激酶的微阵列HTS的初步数据。目标1：将通过与TeleChem(加利福尼亚州桑尼维尔)的财团设计和建造一种化学微阵列。这种微板阵列器将部署一个384针歧管，将NIH库化合物打印到每个玻璃阵列板包含多达38,400个化合物的阵列板(SBS标准)。该仪器的设计目标是在24小时内完成整个NIH储存库的10套印刷，用于10个单独的HTS活动(500万个反应)。目的2：将设计和验证一种阵列板激活剂，用于将亚纳升数量的酶、底物、辅因子和检测器快速输送到复合阵列板。该仪器的设计目标是在30分钟内启动对整个NIH化合物储存库(包含500,000个化合物的20个阵列板)的生化筛选。目标3：构建一个阵列板激活器测试单元并将其交付给美国国立卫生研究院进行测试和验证。该仪器旨在利用微阵列巨大的液体处理能力、用于化合物保存和轻松HTS分析开发的微阵列的小型化，以及多色激光扫描仪的分辨率/灵敏度。该仪器还可用于其他应用：(1)在预涂有固定化蛋白质和结合伙伴的平板上快速打印大型化学库将有助于蛋白质-蛋白质抑制HTS；(2)快速打印100,000或更多在孔板HTS中产生的细胞裂解产物的能力有助于通过夹心ELISA法分析的反相阵列对多种细胞内抗原进行高度多重检测。总体而言，这一仪器允许从NIH储存库的100,000到500,000种化合物中筛选出大量的野生型或重组蛋白靶标，每个靶标的数量都很少。

DESCRIPTION (provided by applicant): Reaction Biology Corporation (Malvern, PA) has developed homogeneous-phase reaction microarrays to serve the NIH Roadmap initiative for nanoliter-scale biochemical high throughput screening (HTS) in drug discovery, large scale IC50 determinations, and selectivity profiling. These nanoliter reactions are 1000 to 10,000-fold smaller than well plate formats currently in use. Preliminary data is presented for microarray HTS against serine proteases, caspases, MMPs, tyrosine kinases, and serine/ threonine kinases. Aim 1: A chemical microarrayer will be designed and built through a Consortium with TeleChem (Sunnyvale, CA). This microplate arrayer will deploy a 384-pin manifold to print NIH library compounds to array-plates containing up to 38,400 compounds per glass array-plate (SBS standard). The design goal for this instrumentation is to achieve the printing of 10 sets of the entire NIH repository in a 24-hr print run to be used for 10 individual HTS campaigns (5 million reactions). Aim 2: An array-plate activator will be designed and validated for the rapid delivery of sub-nanoliter quantities of enzymes, substrates, cofactors, and detectors to the compound array-plates. The design goal for this instrumentation will be to initiate biochemical screens against the entire NIH compound repository (20 array-plates containing 500,000 compounds) in 30 minutes. Aim 3: Construction and delivery of an array-plate activator beta-unit to the NIH for testing and validation. This instrumentation seeks to exploit the tremendous liquid handling power of microarrayers, the miniaturization of microarrays for compound conservation and easy HTS assay development, and the resolution/sensitivity of multicolor laser scanners. Other applications are enabled by this instrumentation: (1) the rapid printing of large chemical libraries on plates pre-coated with an immobilized protein and binding partner would facilitate protein-protein inhibition HTS; (2) the ability to rapidly print 100,000 or more cellular lysates produced in well plate HTS facilitates highly multiplexed detection of numerous intracellular antigens by reverse phase arrays analyzed by sandwich ELISA. Overall, this instrumentation allows a large number of wild type or recombinant protein targets, each obtained in miniscule quantity, to be screened against 100,000 to 500,000 compounds of the NIH repository.