随机光学重构显微镜在外泌体观察中的应用  

Application of stochastic optical reconstruction microscopy in the observation of exosomes

作　　者：马迪[1] 夏仁品[2] 冯志坚[1] 杨筱燕[1] 付珏兰 马各富[3] MA Di1, XIA Renpin2, FENG Zhijian1, YANG Xiaoyan1, FU Juelan1, MA Gefu3(1Department of Kidney Medicine, The Second Afiliated Hospital of Kunming Medical University, Kunming 650500,China; 2Department of Transplantation, The First Afiliated Hospital of Kunming Medical University, Kunming 650032,China; 3Clinical Laboratory, Tianqi Hospital, The Second Affiliated Hospital of Kunming Medical University, Kunming 650500,Chin)

机构地区：[1]昆明医科大学第二附属医院肾脏内科,云南昆明650500 [2]昆明医科大学第一附属医院移植科,云南昆明650032 [3]昆明医科大学第二附属医院石林天奇医院检验科,云南昆明650500

出　　处：《分子影像学杂志》2018年第2期224-228,共5页Journal of Molecular Imaging

摘　　要：目的探讨随机光学重构显微镜(STORM)在外泌体观察中的应用价值,并介绍外泌体在随机光学重构显微镜下的成像原理及方法。方法实验中在肾性甲旁亢患者原代培养的甲状旁腺细胞的培养液上清中加入EXOQUICK-TC外泌体沉淀剂来获取细胞培养上清中的外泌体,将外泌体膜表面的特异性跨膜蛋白CD63作免疫荧光标记后利用随机光学重构显微镜对外泌体行超高分辨率成像并测量外泌体直径。结果实验中随机光学重构显微镜成功的对继发性甲旁亢甲状旁腺细胞原代培养液上清中的外泌体行单分子定位、直径测量及超高分辨率成像。结论因随机光学重构显微镜具有独特的光学特性并突破了光学衍射极限的限制,较传统光学显微镜,能获得20～50 nm的分辨率,可对外泌体行单分子精确定位、直径测量、超高分辨率成像。基于STORM的成像优势,相信STORM及其他超高分辨率成像技术将在外泌体及外泌体参与的生物学过程的研究中发挥重要作用。Objective To investigate the application value of stochastic optical reconstruction microscopy（STORM） in the observation of exosomes. Methods In the experiment, exosomes were extracted by adding EXOQUICK-TC to the culture media supernatants of primarily cultured parathyroid cells from patients with hyperparathyroidism. After the immunofluorescence labeling of the specific transmembrane protein CD63 on the surface of exosomes, STORM was applied to perform ultra-high resolution imaging and diameter measurement of exosomes. Results In the experiment, STORM successfully performed the single molecule positioning, diameter measurement and ultra-high resolution imaging of exosomes extracted from the culture media supernatants of parathyroid cells of patients with hyperparathyroidism. Conclusion Compared to conventional optical microscopy, STORM, with its unique optical characteristics which has broken through the optical diffraction limit, has a resolution of 20-50 nm and can perform single-molecular acute positioning, diameter measurement and ultra-high resolution imaging of exosomes. In view of these outstanding features, we believe that STORM and other ultra-high resolution imaging technologies will play vital roles in the subsequent research on exosomes and exosome-related physiological procedures.

关 键 词：外泌体 随机光学重构显微镜 超高分辨率成像 

分 类 号：R445[医药卫生—影像医学与核医学]