Department of Applied Physics, KTH Royal Institute of Technology/Albanova, 106 91, Stockholm, Sweden.
Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet, 171 77, Solna, Sweden.
Sci Rep. 2017 Oct 18;7(1):13433. doi: 10.1038/s41598-017-13538-2.
Water-window x-ray microscopy allows two- and three-dimensional (2D and 3D) imaging of intact unstained cells in their cryofixed near-native state with unique contrast and high resolution. Present operational biological water-window microscopes are based at synchrotron facilities, which limits their accessibility and integration with complementary methods. Laboratory-source microscopes have had difficulty addressing relevant biological tasks with proper resolution and contrast due to long exposure times and limited up-time. Here we report on laboratory cryo x-ray microscopy with the exposure time, contrast, and reliability to allow for routine high-spatial resolution 3D imaging of intact cells and cell-cell interactions. Stabilization of the laser-plasma source combined with new optics and sample preparation provide high-resolution cell imaging, both in 2D with ten-second exposures and in 3D with twenty-minute tomography. Examples include monitoring of the distribution of carbon-dense vesicles in starving HEK293T cells and imaging the interaction between natural killer cells and target cells.
水窗 X 射线显微镜允许对未经染色的冷冻固定近天然状态的完整细胞进行二维和三维（2D 和 3D）成像，具有独特的对比度和高分辨率。目前运行的生物水窗显微镜基于同步加速器设施，这限制了它们的可及性和与互补方法的集成。由于曝光时间长和正常运行时间有限，实验室光源显微镜难以解决相关的生物学任务，无法实现适当的分辨率和对比度。在这里，我们报告了具有曝光时间、对比度和可靠性的实验室低温 X 射线显微镜，可用于常规的高空间分辨率完整细胞和细胞间相互作用的 3D 成像。激光等离子体源的稳定化结合新的光学器件和样品制备提供了高分辨率的细胞成像，包括使用十秒曝光的 2D 和二十分钟断层扫描的 3D。实例包括监测饥饿的 HEK293T 细胞中碳密集小泡的分布和成像自然杀伤细胞与靶细胞之间的相互作用。
