Eibl Robert H, Moy Vincent T
Department of Physiology and Biophysics, University of Miami School of Medicine, Miami, FL, USA.
Methods Mol Biol. 2005;305:439-50. doi: 10.1385/1-59259-912-5:439.
Cell adhesion receptors are expressed on the surface of cells and can mediate binding to other cells and to the extracellular matrix. Here, we describe in detail the use of atomic force microscopy (AFM)-based force spectroscopy for studying cell detachment forces on living leukocytes. With this technique it is now possible to measure force with resolution down to the level of individual molecules. AFM force spectroscopy is particularly well suited for research in cell adhesion, which has relevance in both the medical and life sciences including immunology, cancer and stem cell research, and human pharmacology. Along with its limitations, we herein, describe how the rupture force of a single complex formed between the integrin receptor leukocyte function-associated antigen (LFA)-1, expressed on the surface of a living leukocyte, and immobilized intercellular adhesion molecule-1 (ICAM-1) was measured. With only minor modifications this protocol can be used to study other adhesion receptors on almost any mammalian cell or bacterial system. This protocol is also suitable for studying single-molecule de-adhesion events in cell-free systems as well as between two living cells.
细胞黏附受体表达于细胞表面，可介导与其他细胞及细胞外基质的结合。在此，我们详细描述基于原子力显微镜（AFM）的力谱技术在研究活白细胞上的细胞脱离力方面的应用。借助该技术，现在能够以低至单个分子水平的分辨率测量力。AFM力谱特别适合细胞黏附研究，这在医学和生命科学领域都具有相关性，包括免疫学、癌症和干细胞研究以及人类药理学。除了其局限性，我们在此描述了如何测量活白细胞表面表达的整合素受体白细胞功能相关抗原（LFA）-1与固定化细胞间黏附分子-1（ICAM-1）之间形成的单个复合物的破裂力。只需进行少量修改，该方案即可用于研究几乎任何哺乳动物细胞或细菌系统上的其他黏附受体。该方案也适用于研究无细胞系统以及两个活细胞之间的单分子去黏附事件。
