High resolution traction force microscopy based on experimental and computational advances
High resolution traction force microscopy based on experimental and computational advances
复制标题
DOI：
10.1529/biophysj.107.113670
复制
发表时间：
2008-01-01
影响因子：
3.4
通讯作者：
Schwarz, Ulrich S.
中科院分区：
文献类型：
--
作者：
Sabass, Benedikt;Gardel, Margaret L.;Schwarz, Ulrich S.
细胞粘附和迁移关键取决于肌动球蛋白产生的力通过粘着斑部位传递到细胞外基质。在这里，我们报告的实验和计算的进步，提高分辨率和可靠性的牵引力显微镜。首先，我们介绍了使用两种不同颜色的纳米珠作为基准标记在聚丙烯酰胺凝胶和解释如何位移场可以计算提取的荧光数据。其次，我们提出了不同的改进标准的方法，从位移场的力重建，这是边界元法，傅立叶变换牵引细胞术，和牵引重建点力。利用大量的数据模拟，我们表明，通过将弹性场分为近场、中场和远场，可以大大提高边界元法的空间分辨率。傅里叶变换牵引细胞术需要相当少的计算机时间，但只有当与维纳滤波或适当的正则化方案相结合时，才能达到相当的分辨率。这两种方法都倾向于低估力，特别是在小的粘连部位。点力牵引重建不受此限制，但仅适用于固定和发育良好的粘连部位。第三，我们结合联合收割机这些进展，并首次重建成纤维细胞牵引与空间分辨率接近1 μ m。
Cell adhesion and migration crucially depend on the transmission of actomyosin-generated forces through sites of focal adhesion to the extracellular matrix. Here we report experimental and computational advances in improving the resolution and reliability of traction force microscopy. First, we introduce the use of two differently colored nanobeads as fiducial markers in polyacrylamide gels and explain how the displacement field can be computationally extracted from the fluorescence data. Second, we present different improvements regarding standard methods for force reconstruction from the displacement field, which are the boundary element method, Fourier-transform traction cytometry, and traction reconstruction with point forces. Using extensive data simulation, we show that the spatial resolution of the boundary element method can be improved considerably by splitting the elastic field into near, intermediate, and far field. Fourier-transform traction cytometry requires considerably less computer time, but can achieve a comparable resolution only when combined with Wiener filtering or appropriate regularization schemes. Both methods tend to underestimate forces, especially at small adhesion sites. Traction reconstruction with point forces does not suffer from this limitation, but is only applicable with stationary and well-developed adhesion sites. Third, we combine these advances and for the first time reconstruct fibroblast traction with a spatial resolution of similar to 1 mu m.