先进成像技术。内吞作用和细胞骨架动力学的超分辨结构光照明显微成像。
ADVANCED IMAGING. Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics.
作者信息
Li Dong, Shao Lin, Chen Bi-Chang, Zhang Xi, Zhang Mingshu, Moses Brian, Milkie Daniel E, Beach Jordan R, Hammer John A, Pasham Mithun, Kirchhausen Tomas, Baird Michelle A, Davidson Michael W, Xu Pingyong, Betzig Eric
机构信息
Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, VA 20147, USA.
Key Laboratory of RNA Biology and Beijing Key Laboratory of Noncoding RNA, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China. College of Life Sciences, Central China Normal University, Wuhan 430079, Hubei, China.
出版信息
Science. 2015 Aug 28;349(6251):aab3500. doi: 10.1126/science.aab3500.
Super-resolution fluorescence microscopy is distinct among nanoscale imaging tools in its ability to image protein dynamics in living cells. Structured illumination microscopy (SIM) stands out in this regard because of its high speed and low illumination intensities, but typically offers only a twofold resolution gain. We extended the resolution of live-cell SIM through two approaches: ultrahigh numerical aperture SIM at 84-nanometer lateral resolution for more than 100 multicolor frames, and nonlinear SIM with patterned activation at 45- to 62-nanometer resolution for approximately 20 to 40 frames. We applied these approaches to image dynamics near the plasma membrane of spatially resolved assemblies of clathrin and caveolin, Rab5a in early endosomes, and α-actinin, often in relationship to cortical actin. In addition, we examined mitochondria, actin, and the Golgi apparatus dynamics in three dimensions.
超分辨率荧光显微镜在纳米级成像工具中独具特色，能够对活细胞中的蛋白质动态进行成像。结构光照明显微镜（SIM）在这方面表现突出，因其速度快且照明强度低，但通常仅能将分辨率提高两倍。我们通过两种方法扩展了活细胞SIM的分辨率：一种是具有84纳米横向分辨率的超高数值孔径SIM，可用于超过100个多色帧的成像；另一种是具有图案激活功能的非线性SIM，分辨率为45至62纳米，可用于大约20至40个帧的成像。我们应用这些方法对网格蛋白和小窝蛋白的空间分辨组装体、早期内体中的Rab5a以及α-辅肌动蛋白（通常与皮质肌动蛋白相关）的质膜附近动态进行成像。此外，我们还对线粒体、肌动蛋白和高尔基体的三维动态进行了研究。
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