School of Biomedical Engineering, Faculty of Engineering, The University of Sydney.
School of Biomedical Engineering, Faculty of Engineering, The University of Sydney; Charles Perkins Centre, The University of Sydney; Heart Research Institute.
J Vis Exp. 2021 Nov 20(177). doi: 10.3791/62490.
A biomembrane force probe (BFP) has recently emerged as a native-cell-surface or in situ dynamic force spectroscopy (DFS) nanotool that can measure single-molecular binding kinetics, assess mechanical properties of ligand-receptor interactions, visualize protein dynamic conformational changes and more excitingly elucidate receptor mediated cell mechanosensing mechanisms. More recently, BFP has been used to measure the spring constant of molecular bonds. This protocol describes the step-by-step procedure to perform molecular spring constant DFS analysis. Specifically, two BFP operation modes are discussed, namely the Bead-Cell and Bead-Bead modes. This protocol focuses on deriving spring constants of the molecular bond and cell from DFS raw data.
生物膜力探针 (BFP) 最近作为一种原生细胞表面或原位动态力谱 (DFS) 纳米工具出现，可测量单分子结合动力学，评估配体-受体相互作用的机械性能，可视化蛋白质动态构象变化，更令人兴奋的是阐明受体介导的细胞机械感觉机制。最近，BFP 已被用于测量分子键的弹性常数。本协议描述了执行分子弹簧常数 DFS 分析的逐步过程。具体来说，讨论了两种 BFP 操作模式，即珠-细胞模式和珠-珠模式。本协议重点从 DFS 原始数据中得出分子键和细胞的弹性常数。
