Noncontact estimation of intercellular breaking force using a femtosecond laser impulse quantified by atomic force microscopy
Noncontact estimation of intercellular breaking force using a femtosecond laser impulse quantified by atomic force microscopy
复制标题
DOI：
10.1073/pnas.1006847108
复制
发表时间：
2011-02-01
影响因子：
11.1
通讯作者：
Ito, Akihiko
中科院分区：
文献类型：
--
作者：
Hosokawa, Yoichiroh;Hagiyama, Man;Ito, Akihiko
当飞秒激光脉冲（fsLP）通过物镜透镜聚焦到培养基中时，产生从激光焦点（O（f））在微米尺寸的空间中传播的冲击力（fsLP-IF）。这种力可以分离单个粘附细胞，而不会造成相当大的细胞损伤。在这项研究中，fsLP-IF反映在原子力显微镜（AFM）悬臂梁的振动运动。基于振动的幅度和O（f）与悬臂梁之间的几何关系，以冲量[N-s]为单位计算O（f）处产生的fsLP-IF。这种冲击力打破了粘附分子介导的细胞间相互作用的方式，依赖于粘附强度，估计通过细胞聚集试验。该力还破坏了链霉亲和素包被的微球和生物素包被的基质之间的相互作用，测量误差约为7%。这些结果表明，fsLP-IF可以用来打破分子间和细胞间的相互作用，并估计粘附强度。fsLP-IF用于打破两种生物学相关培养物中的细胞间接触：接种在内皮细胞单层上的白细胞的共培养物和上皮细胞的极化单层培养物。根据O（f）和粘附界面之间的几何关系计算出的破坏白细胞-内皮和内皮间相互作用所需的脉冲分别为10（-13）和10（-12）N·s。当O（f）处的总冲量定义明确时，fsLP-IF可用于估计在生物学相关条件下以非接触方式破坏细胞间粘附所需的力。
When a femtosecond laser pulse (fsLP) is focused through an objective lens into a culture medium, an impulsive force (fsLP-IF) is generated that propagates from the laser focal point (O(f)) in a micron-sized space. This force can detach individual adherent cells without causing considerable cell damage. In this study, an fsLP-IF was reflected in the vibratory movement of an atomic force microscopy (AFM) cantilever. Based on the magnitude of the vibration and the geometrical relationship between O(f) and the cantilever, the fsLP-IF generated at O(f) was calculated as a unit of impulse [N-s]. This impulsive force broke adhesion molecule-mediated intercellular interactions in a manner that depended on the adhesion strength that was estimated by the cell aggregation assay. The force also broke the interactions between streptavidin-coated microspheres and a biotin-coated substrate with a measurement error of approximately 7%. These results suggest that fsLP-IF can be used to break intermolecular and intercellular interactions and estimate the adhesion strength. The fsLP-IF was used to break intercellular contacts in two biologically relevant cultures: a coculture of leukocytes seeded over on an endothelial cell monolayer, and a polarized monolayer culture of epithelial cells. The impulses needed to break leukocyte-endothelial and interepithelial interactions, which were calculated based on the geometrical relationship between O(f) and the adhesive interface, were on the order of 10(-13) and 10(-12) N-s, respectively. When the total impulse at O(f) is well-defined, fsLP-IF can be used to estimate the force required to break intercellular adhesions in a noncontact manner under biologically relevant conditions.