Hearty Stephen, Leonard Paul, Ma Hui, O'Kennedy Richard
School of Biotechnology, Dublin City University, Dublin, Ireland.
National Centre for Sensor Research, Dublin City University, Dublin, Ireland.
Methods Mol Biol. 2018;1827:421-455. doi: 10.1007/978-1-4939-8648-4_22.
Surface plasmon resonance (SPR) is now widely embraced as a technology for monitoring a diverse range of protein-protein interactions and is considered almost de rigueur for characterizing antibody-antigen interactions. The technique obviates the need to label either of the interacting species, and the binding event is visualized in real time. Thus, it is ideally suited for screening crude, unpurified antibody samples that dominate early candidate panels following antibody selection campaigns. SPR returns not only concentration and affinity data but when used correctly can resolve the discrete component kinetic parameters (association and dissociation rate constants) of the affinity interaction. Herein, we outline some SPR-based generic antibody screening configurations and methodologies in the context of expediting data-rich ranking of candidate antibody panels and ensuring that antibodies with the optimal kinetic binding characteristics are reliably identified.
表面等离子体共振(SPR)如今已被广泛应用,作为一种监测多种蛋白质-蛋白质相互作用的技术,几乎被视为表征抗体-抗原相互作用的必备手段。该技术无需对任何一种相互作用的分子进行标记,并且能实时观察到结合事件。因此,它非常适合筛选粗制、未纯化的抗体样品,这些样品在抗体筛选活动后的早期候选库中占主导地位。SPR不仅能返回浓度和亲和力数据,而且如果使用得当,还能解析亲和力相互作用的离散组分动力学参数(结合和解离速率常数)。在此,我们概述了一些基于SPR的通用抗体筛选配置和方法,旨在加快对候选抗体库进行富含数据的排名,并确保可靠地鉴定出具有最佳动力学结合特性的抗体。