Friedrichs Jens, Werner Carsten, Müller Daniel J
Leibniz Institute of Polymer Research Dresden, Institute for Biofunctional Polymer Materials, Dresden, Germany.
Methods Mol Biol. 2013;1046:19-37. doi: 10.1007/978-1-62703-538-5_2.
Atomic force microscopy (AFM)-based single-cell force spectroscopy (SCFS) enables the quantitative study of cell adhesion under physiological conditions. SCFS probes adhesive interactions of single living cells with substrates such as extracellular matrix (ECM) proteins and other cells. Here, we present a protocol to quantitatively study the adhesion of HeLa cells to covalently immobilized fibronectin and Matrigel™ using SCFS. We describe procedures for (a) functionalization of AFM cantilevers, (b) preparation of maleic anhydride copolymer thin films, (c) covalent immobilization of ECM proteins on the thin films, (d) cell handling and attachment to the AFM cantilever, and (e) measurement of adhesion forces. The protocol can be easily modified for other cell types and substrate proteins.
基于原子力显微镜（AFM）的单细胞力谱（SCFS）能够在生理条件下对细胞黏附进行定量研究。SCFS可探测单个活细胞与诸如细胞外基质（ECM）蛋白和其他细胞等底物之间的黏附相互作用。在此，我们展示了一种使用SCFS定量研究HeLa细胞与共价固定的纤连蛋白和基质胶™黏附的方案。我们描述了以下步骤：（a）AFM悬臂的功能化，（b）马来酸酐共聚物薄膜的制备，（c）ECM蛋白在薄膜上的共价固定，（d）细胞处理及与AFM悬臂的附着，以及（e）黏附力的测量。该方案可轻松修改以适用于其他细胞类型和底物蛋白。
