a multi-functional imaging approach to high-content protein interaction screening高含量的蛋白质相互作用筛选多功能成像方法.pdfVIP

A Multi-Functional Imaging Approach to High-Content Protein Interaction Screening 1 1 1 1¤ 1 Daniel R. Matthews , Gilbert O. Fruhwirth , Gregory Weitsman , Leo M. Carlin , Enyinnaya Ofo , 1 1,2 2 1,2 1 Melanie Keppler , Paul R. Barber , Iain D. C. Tullis , Borivoj Vojnovic , Tony Ng , Simon M. Ameer- Beg1* 1 Division of Cancer Studies, Randall Division of Cell and Molecular Biophysics, King’s College London, London, United Kingdom, 2 Gray Institute for Radiation Oncology and Biology, Department of Oncology, University of Oxford, Oxford, United Kingdom Abstract Functional imaging can provide a level of quantification that is not possible in what might be termed traditional high- content screening. This is due to the fact that the current state-of-the-art high-content screening systems take the approach of scaling-up single cell assays, and are therefore based on essentially pictorial measures as assay indicators. Such phenotypic analyses have become extremely sophisticated, advancing screening enormously, but this approach can still be somewhat subjective. We describe the development, and validation, of a prototype high-content screening platform that combines steady-state fluorescence anisotropy imaging with fluorescence lifetime imaging (FLIM). This functional approach allows objective, quantitative screening of small molecule libraries in protein-protein interaction assays. We discuss the development of the instrumentation, the process by which information on fluorescence resonance energ