Assay development and screening of a serine/threonine kinase in an on-chip mode using caliper nanofluidics technology
Assay development and screening of a serine/threonine kinase in an on-chip mode using caliper nanofluidics technology
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DOI：
10.1177/1087057106286653
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发表时间：
2006-06-01
影响因子：
--
通讯作者：
Scheer, Alexander
中科院分区：
文献类型：
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作者：
Perrin, Dominique;Fremaux, Christele;Scheer, Alexander
激酶是药物发现的关键靶点。在一般的筛选领域中，特别是在激酶领域中，由于效率和成本的考虑，基于放射性的测定往往被替代的、主要是基于荧光的测定所取代。今天，限制因素很少是可以获得的数据点的数量，而是数据的质量，酶的可用性和成本。在这篇文章中，作者描述了一种基于荧光产物和底物的电泳分离的激酶筛选试验的开发，该试验使用基于卡尺的纳米流体环境在芯片上孵育模式下进行。作者介绍了筛选一组32，000种化合物的结果，以及在过滤试验中获得的确认数据。此外，他们还对芯片上和芯片外纳米流体筛选模式进行了小规模比较。在他们手中，片上模式下的屏幕具有高精度的特点，很可能是由于没有液体移液;在独立的测定形式（即过滤）中具有出色的确认率（62%）;以及良好的灵敏度。这项研究导致了4个新的化学系列的抑制剂的鉴定。
Kinases are key targets for drug discovery. In the field of screening in general and especially in the kinase area, because of considerations of efficiency and cost, radioactivity-based assays tend to be replaced by alternative, mostly fluorescence-based, assays. Today, the limiting factor is rarely the number of data points that can be obtained but rather the quality of the data, enzyme availability, and cost. In this article, the authors describe the development of an assay for a kinase screen based on the electrophoretic separation of fluorescent product and substrate using a Caliper-based nanofluidics environment in on-chip incubation mode. The authors present the results of screening a focused set of 32,000 compounds together with confirmation data obtained in a filtration assay. In addition, they have made a small-scale comparison between the on-chip and off-chip nanofluidics screening modes. In their hands, the screen in on-chip mode is characterized by high precision most likely due to the absence of liquid pipetting; an excellent confirmation rate (62%) in an independent assay format, namely, filtration; and good sensitivity. This study led to the identification of 4 novel chemical series of inhibitors.