Department of Chemical Engineering, Stanford University, Stanford, CA 94305, United States.
Stanford-SLAC CryoEM Initiative, Stanford, CA 94305, United States.
J Struct Biol. 2021 Dec;213(4):107791. doi: 10.1016/j.jsb.2021.107791. Epub 2021 Sep 11.
Cryo-electron tomography is the highest resolution tool available for structural analysis of macromolecular complexes within their native cellular environments. At present, data acquisition suffers from low throughput, in part due to the low probability of positioning a cell such that the subcellular structure of interest is on a region of the electron microscopy (EM) grid that is suitable for imaging. Here, we photo-micropatterned EM grids to optimally position endothelial cells so as to enable high-throughput imaging of cell-cell contacts. Lattice micropatterned grids increased the average distance between intercellular contacts and thicker cell nuclei such that the regions of interest were sufficiently thin for direct imaging. We observed a diverse array of membranous and cytoskeletal structures at intercellular contacts, demonstrating the utility of this technique in enhancing the rate of data acquisition for cellular cryo-electron tomography studies.
冷冻电子断层扫描是用于分析天然细胞环境中大分子复合物结构的最高分辨率工具。目前，数据采集的速度较慢，部分原因是将细胞定位在电子显微镜（EM）网格上适合成像的亚细胞结构区域的可能性较低。在这里，我们对 EM 网格进行了光微图案化处理，以最佳方式定位内皮细胞，从而实现细胞-细胞接触的高通量成像。晶格微图案化网格增加了细胞间接触和较厚细胞核之间的平均距离，使得感兴趣的区域足够薄，可以直接成像。我们观察到细胞间接触处存在各种各样的膜和细胞骨架结构，证明了该技术在提高细胞冷冻电子断层扫描研究的数据采集速度方面的实用性。
