一种用于线粒体受激辐射损耗超分辨成像的新型探针  

Study on a novel probe for stimulated emission depletion Super-resolution Imaging of Mitochondria

作　　者：张佳 Samanta Soham 王佳林[1] 王璐玮 杨志刚 严伟[1] 屈军乐[1] Zhang Jia;Samanta Soham;Wang Jia-Lin;Wang Lu-Wei;Yang Zhi-Gang;Yan Wei;Qu Jun-Le(Key Laboratory of Optoelectronic Devices and Systems of Ministry of Education and Guangdong Province,College of Physics and Optoelectronic Engineering,Shenzhen University,Shenzhen 518060,China)

机构地区：[1]深圳大学物理与光电工程学院,光电子器件与系统教育部/广东省重点实验室,深圳518060

出　　处：《物理学报》2020年第16期304-312,共9页Acta Physica Sinica

基　　金：国家重点基础研究发展计划(批准号:2017YFA0700500);国家自然科学基金(批准号:61975127,61525503,81727804);广东省高等学校科技创新项目(批准号:2016KCXTD007);广东省自然科学基金(批准号:2020A1515010679);深圳市基础研究项目(批准号:JCYJ2018030512304883,JCYJ20170818100153423,JCYJ20170412105003520)资助的课题.

摘　　要：受激辐射损耗(stimulated emission depletion,STED)显微技术通过巧妙的光学设计,利用纯光学的方法突破了光学衍射极限,空间分辨率达到纳米量级,并保留了荧光显微的许多优点.然而,高的损耗光强度限制了STED显微技术的广泛应用,尤其在活细胞成像方面.本文找到了一种新型的具有良好线粒体靶向性的STED探针,具有较强的抗光漂白特性和较低的饱和擦除光强度3.5 mW(1.1 MW·cm–2),利用该探针最高可获得62 nm的空间分辨率,为活细胞线粒体STED超分辨成像提供了新的手段.Optical microscopy has the advantages of real-time,non-invasive,tomography,three-dimensional imaging and living imaging.However,its spatial resolution cannot exceed half wavelength due to the existence of optical diffraction limit,which limits the development of optical microscopy.The primary task of super-resolution imaging is to break the diffraction limit and improve the resolution of optical microscopy for study of subcellular structure.Many kinds of super-resolution imaging technologies have been reported,among which the stimulated emission depletion(STED)microscopy is the earliest imaging technology to break the optical diffraction limit at present.STED microscopy can achieve nanometer-scale spatial resolution by breaking the optical diffraction limit with pure optical methods and a clever optical design.However,the application of STED microscopy in biomedicine,especially in live cell imaging is limited by high illumination power of STED light.In this paper,a new type of STED probe has been developed.The spectral analysis results show that the peak of the excitation and emission spectrum of this probe is as far as 122 nm away from each other,which is very suitable for the study of STED super-resolution because of its long stokes redshift.After colocalization with commercial mitochondrial dyes,it was found that the probe had a higher localization coefficient with commercial dyes and could be well positioned on mitochondrial organelles.At the same time,it was found that strong mitochondrial signal could be detected with low-power excitation light(only 1μW in the experiment),and can get higher resolution of 62 nm under the STED light with 39.5 mW.The result of measuring the transverse resolution obtained by STED light under different power shows that the saturated light power of the probe is 3.5 mW(1.1 MW·cm–2).Through the anti-bleaching testing,the probe still has a strong fluorescence intensity after more than 300 times of high power light irradiation,which indicates that the probe has a strong anti-blea

关 键 词：受激辐射损耗显微 超分辨成像 荧光探针 线粒体 活细胞成像 

分 类 号：O657.3[理学—分析化学] R313[理学—化学]