Nierode Gregory, Kwon Paul S, Dordick Jonathan S, Kwon Seok-Joon
Department of Chemical and Biological Engineering and Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY 12180, USA.
J Microbiol Biotechnol. 2016 Feb;26(2):213-25. doi: 10.4014/jmb.1508.08007.
To reduce attrition in drug development, it is crucial to consider the development and implementation of translational phenotypic assays as well as decipher diverse molecular mechanisms of action for new molecular entities. High-throughput fluorescence and confocal microscopes with advanced analysis software have simplified the simultaneous identification and quantification of various cellular processes through what is now referred to as highcontent screening (HCS). HCS permits automated identification of modifiers of accessible and biologically relevant targets and can thus be used to detect gene interactions or identify toxic pathways of drug candidates to improve drug discovery and development processes. In this review, we summarize several HCS-compatible, biochemical, and molecular biology-driven assays, including immunohistochemistry, RNAi, reporter gene assay, CRISPR-Cas9 system, and protein-protein interactions to assess a variety of cellular processes, including proliferation, morphological changes, protein expression, localization, post-translational modifications, and protein-protein interactions. These cell-based assay methods can be applied to not only 2D cell culture but also 3D cell culture systems in a high-throughput manner.
为了减少药物研发中的损耗，考虑转化型表型分析的开发与实施以及解读新分子实体的多种分子作用机制至关重要。配备先进分析软件的高通量荧光显微镜和共聚焦显微镜，通过现在所谓的高内涵筛选（HCS）简化了对各种细胞过程的同时识别和定量。HCS允许自动识别可及且生物学相关靶点的调节剂，因此可用于检测基因相互作用或识别候选药物的毒性途径，以改善药物发现和研发过程。在本综述中，我们总结了几种与HCS兼容的、由生化和分子生物学驱动的分析方法，包括免疫组织化学、RNA干扰、报告基因分析、CRISPR-Cas9系统以及蛋白质-蛋白质相互作用，以评估包括增殖、形态变化、蛋白质表达、定位、翻译后修饰和蛋白质-蛋白质相互作用在内的多种细胞过程。这些基于细胞的分析方法不仅可以高通量方式应用于二维细胞培养，还可应用于三维细胞培养系统。
