Paduch Marcin, Kossiakoff Anthony A
Department of Biochemistry and Molecular Biology, The University of Chicago, KCBD Rm.3240G, 900 East 57th Street, Chicago, IL, 60637, USA.
Institute for Biophysical Dynamics, The University of Chicago, Chicago, IL, 60637, USA.
Methods Mol Biol. 2017;1575:93-119. doi: 10.1007/978-1-4939-6857-2_6.
Phage display is commonly used to identify and isolate binders from large combinatorial libraries. Here we present phage selection protocols enabling generation of synthetic antibodies capable of recognizing multiprotein complexes and conformational states. The procedure describes stages of the experiment design, optimization, and screening, as well as provides the framework for building downstream assays with an end goal of isolating bioactive antibodies for future therapeutic use. The methods described are also applicable to screening directly on cells and can be ported to other in vitro directed evolution systems utilizing non-immunoglobulin scaffolds.
噬菌体展示常用于从大型组合文库中识别和分离结合物。在此，我们介绍噬菌体筛选方案，该方案能够生成能够识别多蛋白复合物和构象状态的合成抗体。该程序描述了实验设计、优化和筛选的各个阶段，并为构建下游分析提供了框架，最终目标是分离出具有生物活性的抗体以供未来治疗使用。所描述的方法也适用于直接在细胞上进行筛选，并且可以移植到利用非免疫球蛋白支架的其他体外定向进化系统中。
