Studies on cell movement and cvtoskeleton by atomic force microscopy

原子力显微镜（AFM）没有透镜，而是简单地在样品表面上扫描尖锐的探针，以从微米尺度到原子尺度的高分辨率提供样品的形貌图像。在本研究中，我们将原子力显微镜应用于活体状态下培养的动物细胞的三维观察，由于球形细胞在扫描过程中容易从基底表面脱离，因此我们使用了牢固附着在基底上的扁平细胞。为了使力引起的细胞损伤最小化，将成像力调节至最弱值。因此，活细胞的接触模式AFM提供了关于细胞过程（尖峰状过程、板状伪足等）的形状的精确信息。在细胞边缘。在细胞膜下的细胞骨架轮廓在细胞上表面也清晰可见，然后我们成功地获得了一个以上的时间间隔为2-4 min的连续AFM图像的活细胞的一个多小时，通过使用一个流体室系统。这些AFM图像的一系列是有用的检查有关亚细胞骨架元素的细胞过程的运动。由连续AFM图像产生的延时电影也验证了细胞动力学的真实性，并将AFM图像与荧光显微镜或扫描电子显微镜拍摄的图像进行了比较。这些数据表明，肌动蛋白积累的颗粒隆起的领导过程中存在。

The atomic force microscope (AFM) has no lens but simply scans a sharp probing tip over a sample surface, providing topographic images of samples at high resolutions from the micrometer scale to the atomic scale. In this study, we applied AFM to the three-dimensional observation of cultured animal cells in a living state.We used flat cells attached firmly to the substrate, becauce spherical cells were easily detached from the substrate surface during scanning. To minimize force-induced damages to the cell, imaging force was adjusted to the weakest value. Thus, contact mode AFM of the living cells provided precise information on the shape of cellular processes (spike-like processes, lamellipodia, etc.) at the cellular margin. The contour of cytoskeletal elements just beneath the cell membrane was also clearly observable on the upper surface of the cells.We then succeeded in obtaining continuously AFM images of living cells for over one hour at time intervals of 2-4 min by using a fluid chamber system. A series of these AFM images were useful for examining the movements of cellular processes in relation to subcellular cytoskeletal elements. Time-lapse movies produced by sequential AFM images also verified the reality of the cellular dynamics.Wefinally compared the AFM images with images taken by fluorescent microscopy, or scanning electron microscopy. The data indicated the presence of actin accumulation in granular elevations on the leading processes.