SENSITIVE FORCE TECHNIQUE TO PROBE MOLECULAR ADHESION AND STRUCTURAL LINKAGES AT BIOLOGICAL INTERFACES
SENSITIVE FORCE TECHNIQUE TO PROBE MOLECULAR ADHESION AND STRUCTURAL LINKAGES AT BIOLOGICAL INTERFACES
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DOI：
10.1016/s0006-3495(95)80441-8
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发表时间：
1995-06-01
影响因子：
3.4
通讯作者：
MERKEL, R
中科院分区：
文献类型：
--
作者：
EVANS, E;RITCHIE, K;MERKEL, R
粘附和细胞骨架结构与生物细胞功能密切相关。即使存在大量的生物和生化数据，但在分子水平上，人们对细胞间附着所涉及的物理机制或粘附对材料结构的后果知之甚少。为了揭示软生物界面的物理行为，我们结合了超灵敏传感器和反射干涉显微镜，对微小力下探针与测试表面接触的亚微观位移进行成像。该传感器是一个细胞大小的膜囊，通过微量吸管吸力加压，其中在张力下垂直于膜的位移与所施加的力成正比。张力的压力控制通过从 0.01 pN 到共价键强度（类似于 1000 pN）的力范围将操作灵敏度调整超过四个数量级！作为表面探针，微小的珠子通过生化方法粘合到传感器上，其具有与测试表面无关的紧密结合的配体。根据珠子反射光产生的干涉条纹图案，可以将探头在施加力下的运动分辨率降低到类似于 5 nm 的精度。通过这种布置，我们表明可以在由结构波动设定的位移分辨率下测量细胞表面的局部机械柔量。当需要时，第二个配体稀疏地结合到探针上，以粘附到测试表面上的特定受体。我们证明，在低换能器刚度下监测探针位置的波动可以增强对分子粘附和细胞骨架结构激活的检测。随后加载附件，测试在整个力驱动的分离过程中受体-基底连接的机械响应。
Adhesion and cytoskeletal structure are intimately related in biological cell function. Even with the vast amount of biological and biochemical data that exist, little is known at the molecular level about physical mechanisms involved in attachments between cells or about consequences of adhesion on material structure. To expose physical actions at soft biological interfaces, we have combined an ultrasensitive transducer and reflection interference microscopy to image submicroscopic displacements of probe contact with a test surface under minuscule forces. The transducer is a cell-size membrane capsule pressurized by micropipette suction where displacement normal to the membrane under tension is proportional to the applied force. Pressure control of the tension tunes the sensitivity in operation over four orders of magnitude through a range of force from 0.01 pN up to the strength of covalent bonds (similar to 1000 pN)! As the surface probe, a microscopic bead is biochemically glued to the transducer with a densely-bound ligand that is indifferent to the test surface. Movements of the probe under applied force are resolved down to an accuracy of similar to 5 nm from the interference fringe pattern created by light reflected from the bead. With this arrangement, we show that local mechanical compliance of a cell surface can be measured at a displacement resolution set by structural fluctuations. When desired, a second ligand is bound sparsely to the probe for focal adhesion to specific receptors in the test surface. We demonstrate that monitoring fluctuations in probe position at low transducer stiffness enhances detection of molecular adhesion and activation of cytoskeletal structure. Subsequent loading of an attachment tests mechanical response of the receptor-substrate linkage throughout the force-driven process of detachment.