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超分辨率成像荧光探针材料应用进展

刘志贺 吴长锋

刘志贺, 吴长锋. 超分辨率成像荧光探针材料应用进展[J]. 中国光学, 2018, 11(3): 344-362. doi: 10.3788/CO.20181103.0344
引用本文: 刘志贺, 吴长锋. 超分辨率成像荧光探针材料应用进展[J]. 中国光学, 2018, 11(3): 344-362. doi: 10.3788/CO.20181103.0344
LIU Zhi-he, WU Chang-feng. Advances in application of materials of super-resolution imaging fluorescent probe[J]. Chinese Optics, 2018, 11(3): 344-362. doi: 10.3788/CO.20181103.0344
Citation: LIU Zhi-he, WU Chang-feng. Advances in application of materials of super-resolution imaging fluorescent probe[J]. Chinese Optics, 2018, 11(3): 344-362. doi: 10.3788/CO.20181103.0344

超分辨率成像荧光探针材料应用进展

doi: 10.3788/CO.20181103.0344
基金项目: 

国家自然科学基金 Grant No.61335001

国家自然科学基金 Grant No.81771930

深圳市科技创新委员会基金 Grant No.JCYJ20170307110157501

详细信息
    作者简介:

    刘志贺(1992—),男,辽宁朝阳人,博士研究生,2016年获得吉林大学硕士学位,主要从事超分辨荧光成像及荧光探针方面的研究。E-mail:liuzhihe002@163.com

    吴长锋(1976—),男,山东德州人,博士,教授,博士生导师。主要从事半导体聚合物点(Polymer Dot)作为新型生物荧光探针的研究,拓展聚合物点在超分辨细胞成像、活体成像、生物传感器、光动力学治疗等领域的应用。E-mail:wucf@sustc.edu.cn

  • 中图分类号: R318.51

Advances in application of materials of super-resolution imaging fluorescent probe

Funds: 

the National Natural Science Foundation of China Grant No.61335001

the National Natural Science Foundation of China Grant No.81771930

Shenzhen Science and Technology Innovation Commission Grant No.JCYJ20170307110157501

More Information
  • 摘要: 为了进一步认知复杂环境中的细胞生物学过程,研究人员发展了各种各样的生物成像技术。在这些技术中,生物荧光成像因简单的成像条件以及对生物样品的相容性而得到了广泛的发展。然而,传统的荧光成像技术受到了光学衍射极限的限制,无法分辨低于200 nm的空间结构,阻碍了对亚细胞结构的生物学过程研究。超分辨荧光显微镜技术突破了传统光学衍射对成像分辨率的限制,能够获取纳米尺度的细胞动态过程。除了对传统的宽场荧光显微镜框架的改进及升级改造之外,目前典型的超分辨成像显微镜技术通常依赖于荧光探针材料的光物理性质。常用的荧光探针材料包括荧光蛋白、有机荧光分子和纳米荧光材料等。本文介绍了几种主流的超分辨荧光显微成像技术并总结了已经成功应用到超分辨生物荧光成像中的荧光探针材料的应用进展。
  • 图  1  不同生物成像技术空间分辨率对比图

    Figure  1.  Spatial resolution comparison of different biological imaging techniques

    图  2  STED超分辨显微镜原理示意图[25]

    Figure  2.  Basic light-pathway diagram and the principle of STED microscopy[25]

    图  3  红光发射的罗丹明荧光分子结构式

    Figure  3.  Chemical structures of red-emitting rhodamine fluorophores

    图  4  用于STED荧光显微镜的荧光分子1及其衍生物结构式

    Figure  4.  Chemical structures of fluorescent molecular 1 and derivatives used in STED microscopy

    图  5  荧光分子2和3的化学结构式及在活细胞中的正交反应示意图

    Figure  5.  Chemical structures of fluorescent moleculars 2 and 3 and the bioorthogonal reaction in live cells

    图  6  荧光分子4~7的化学结构式

    Figure  6.  Chemical structures of fluorescent moleculars 4~7

    图  7  光激活定位显微镜和随机光学重构显微镜工作原理及亚细胞结构超分辨图(左侧:PALM及COS-7细胞溶酶体超分辨图像;右侧:STORM原理图及亚细胞结构单色、多色超分辨图像)[14, 17]

    Figure  7.  Working principle of Photoactivation Localization Microscopy, Stochastic Optical Reconstruction Microscopy and subcellular structure super resolution images(Left: PALM and COS-7 cell lysosomal superresolution images; right:STORM schematic and subcellular structure monochrome, multicolor super-resolution images)[14, 17]

    图  8  荧光分子8~10的化学结构式

    Figure  8.  Chemical structures of fluorescent moleculars 8~10

    图  9  罗丹明衍生物(11~21)光诱导转换过程分子结构式

    Figure  9.  Photoinduced reversible switching process of rhodamine derivatives(11~21)

    图  10  Carbofluoresceins,Carborhodamines110的分子结构式

    Figure  10.  Chemical structures of carbofluoresceins, carborhodamines110(28)

    图  11  分子25~32开关过程的化学结构式以及通过荧光分子31实现的细胞超分辨率图像;紫外光照射下分子34的光开关转换过程[52-53]

    Figure  11.  Chemical structures and the switching process of 25~32 and superresolution images obtained with 31 in live cells. Photoswitchable process of 34 under irradiation with UV/visible light[52-53]

    图  12  (a~c)3种不同DNA结构的三色STORM图像(Alexa405/Cy5, Cy2/Cy5和Cy3/Cy5标记;405 nm, 457 nm和532 nm激光激发);(d)Alexa647/Cy3标记的BC-S-1细胞内网格蛋白凹坑(CCPs)的三维STORM图像[54-55]

    Figure  12.  (a~c)Three-color STORM images of three different DNA constructs(labelled with Alexa405/Cy5, Cy2/Cy5, and Cy3/Cy5, activated using 405, 457 and 532 nm laser, respectively); (d)3D STORM image of clathrin-coated pits(CCPs) stained with the Alexa647/Cy3 in BS-C-1 cells[54-55]

    图  13  花青染料(35~40)分子式和假设的闪烁机制

    Figure  13.  Chemical structures of cyanine dyes(35~40) and the proposed photoblinking mechanism

    图  14  STORM荧光探针41~48结构式及细胞微管的超分辨图像[62];Si-罗丹明荧光分子(49~55)衍生物以及分子54在合适pH环境中闪烁机制的模型及相应的化学结构式[65]; ATTO655标记的CCR5在水和重水不同环境下,分子56和57在光照时互相转化过程以及亚细胞结构图像[66]

    Figure  14.  Chemical structures of fluorescent dyes(41-48) used in STORM[62]; Chemical structures of 49-55 and proposed photoblinking mechanism of 54 in a suitable pH environment[65]; Photoswitching between 56 and 57 upon photoirradiation in the presence of oxygen and ATTO655-labelled CCR5 imaging of live cells in H2O and D2O[66]

    图  15  SOFI成像原理示意图及使用Qdots标记的细胞微管SOFI图像[19, 67, 72]

    Figure  15.  Principle of SOFI imaging and SOFI images of cell microtubule labeled with Qdots[19, 67, 72]

    图  16  聚合物荧光探针分子58~59化学结构式,荧光闪烁特性和单色细胞微管标记图像[83]

    Figure  16.  Chemical structure and photoblinking property of fluorescent polymer probe molecules(58~59), single-color microtubule images labelled with polymer dots[83]

    图  17  聚合物荧光探针分子61~62化学结构式,荧光闪烁特性和细胞微管多色标记图像[84]

    Figure  17.  Chemical structure and photoblinking property of fluorescent polymer probe molecules(61~62), multi-color microtubule images labelled with polymer dots[84]

    图  18  SIM/SSIM原理图, 宽场显微镜和SIM显微镜下GFP标记的COS-7细胞线粒体结构图

    Figure  18.  Principlephotos of SIM and SSIM. GFP-stained mitochondria in COS-7 cells imaged with conventional wide-field and structured illumination microscopy

  • [1] FERRARI M. Cancer nanotechnology:opportunities and challenges[J]. Nature Reviews:Cancer, 2005, 5(3):161-171. doi:  10.1038/nrc1566
    [2] NIE S, XING Y, KIM G J, et al.. Nanotechnology applications in cancer[J]. Annual Review of Biomedical Engineering, 2007, 9:257-288. doi:  10.1146/annurev.bioeng.9.060906.152025
    [3] BEN N G GIEPMANS, STEPHEN R ADAMS, MARK H ELLISMAN, et al.. The fluorescent toolbox for assessing protein location and function[J]. Science, 2006, 312(217):224. http://cn.bing.com/academic/profile?id=cc996e1880426cdd60a036f11b646509&encoded=0&v=paper_preview&mkt=zh-cn
    [4] LI G W, XIE X S. Central dogma at the single-molecule level in living cells[J]. Nature, 2011, 475(7356):308-315. doi:  10.1038/nature10315
    [5] XIE X. S, YU J, YANG W Y. Living cells as test tubes[J]. Science, 2006, 312:228-230. doi:  10.1126/science.1127566
    [6] ABBE E. Beiträge zur Theorie des Mikroskops und der mikroskopischen Wahrnehmung[J]. Archiv Für Mikroskopische Anatomie, 1873, 9(1):413-418. doi:  10.1007/BF02956173
    [7] KLEIN T, PROPPERT S, SAUER M. Eight years of single-molecule localization microscopy[J]. Histochemistry and Cell Biology, 2014, 141(6):561-575. doi:  10.1007/s00418-014-1184-3
    [8] STEFAN W. H, WICHMANN J. Breaking the diffraction resolution limit by stimulated emission stimulatedemission depletion fluorescence microscopy[J]. Optics Letters, 1994, 19(11):780-782. doi:  10.1364/OL.19.000780
    [9] GAEL M, REBECCA M, BIRKA H, et al.. Fast STED microscopy with continuous wave fiber lasers[J]. Optics Express, 2010, 18(2):1302-1309. doi:  10.1364/OE.18.001302
    [10] SUSANNE S, THORSTEN S, RITTWEGER E, et al.. STED nanoscopy with mass-produced laser diodes[J]. Optics Express, 2011, 19(9):8066-8072. doi:  10.1364/OE.19.008066
    [11]