Wu Zheng-Xing, Xia Sheng, Xu Liang, Bai Li, Xu Tao
Institute of Biophysics and Biochemistry, Huazhong University of Science and Technology, Wuhan 430074, China.
Sheng Wu Hua Xue Yu Sheng Wu Wu Li Xue Bao (Shanghai). 2003 Apr;35(4):381-6.
Fluorescent labeling and dynamic imaging of secretory vesicles are new powerful means to study the mechanisms of protein and membrane trafficking. The large dense-core granules in PC12 cells were labeled by transfecting EGFP-hpNPY chimera and imaged with epi-fluorescence and evanescent field (EF) fluorescence simultaneously. Under epi-fluorescence illumination, the cells exhibited nearly uniform fluorescence, however, EF-fluorescence excitation revealed distinct fluorescent spots corresponding to GFP-labeled granules. The trafficking, docking and fusing with plasma membrane of individual granule in cultured PC12 cells were observed directly by EF fluorescence imaging.
分泌囊泡的荧光标记和动态成像为研究蛋白质和膜运输机制提供了新的有力手段。通过转染EGFP-hpNPY嵌合体对PC12细胞中的大致密核心颗粒进行标记，并同时利用落射荧光和倏逝场（EF）荧光进行成像。在落射荧光照明下，细胞呈现出几乎均匀的荧光，然而，EF荧光激发显示出与GFP标记颗粒相对应的明显荧光斑点。通过EF荧光成像直接观察了培养的PC12细胞中单个颗粒与质膜的运输、对接和融合。
