在表达 GFP 的活体多细胞生物中进行纳米显微镜检查。
Nanoscopy in a living multicellular organism expressing GFP.
机构信息
Department of NanoBiophotonics, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany.
出版信息
Biophys J. 2011 Jun 22;100(12):L63-5. doi: 10.1016/j.bpj.2011.05.020.
Abstract
We report superresolution fluorescence microscopy in an intact living organism, namely Caenorhabditis elegans nematodes expressing green fluorescent protein (GFP)-fusion proteins. We also superresolve, by stimulated emission depletion (STED) microscopy, living cultured cells, demonstrating that STED microscopy with GFP can be widely applied. STED with GFP can be performed with both pulsed and continuous-wave lasers spanning a wide wavelength range from at least 556-592 nm. Acquiring subdiffraction resolution images within seconds enables the recording of movies revealing structural dynamics. These results demonstrate that numerous microscopy studies of live samples employing GFP as the marker can be performed at subdiffraction resolution.
摘要
我们在一个完整的活体生物中报告了超分辨率荧光显微镜，即表达绿色荧光蛋白（GFP）融合蛋白的秀丽隐杆线虫。我们还通过受激发射损耗（STED）显微镜对活培养细胞进行了超分辨，证明了 GFP 的 STED 显微镜可以广泛应用。 GFP 的 STED 可以使用从至少 556-592nm 的宽波长范围的脉冲和连续波激光器进行。在几秒钟内获取亚衍射分辨率的图像，可以记录显示结构动力学的电影。这些结果表明，许多使用 GFP 作为标记的活样本的显微镜研究可以在亚衍射分辨率下进行。
相似文献
1
Nanoscopy in a living multicellular organism expressing GFP.在表达 GFP 的活体多细胞生物中进行纳米显微镜检查。
Biophys J. 2011 Jun 22;100(12):L63-5. doi: 10.1016/j.bpj.2011.05.020.
2
Comparing video-rate STED nanoscopy and confocal microscopy of living neurons.比较活神经元的视频速率 STED 纳米显微镜和共聚焦显微镜。
J Biophotonics. 2010 Jul;3(7):417-24. doi: 10.1002/jbio.201000038.
3
Dual-label STED nanoscopy of living cells using photochromism.利用光致变色现象对活细胞进行双标记 STED 纳米显微镜检测。
Nano Lett. 2011 Sep 14;11(9):3970-3. doi: 10.1021/nl202290w. Epub 2011 Aug 8.
4
Coaligned dual-channel STED nanoscopy and molecular diffusion analysis at 20 nm resolution.20nm 分辨率下共定位双通道 STED 纳米显微镜与分子扩散分析
Biophys J. 2013 Jul 2;105(1):L01-3. doi: 10.1016/j.bpj.2013.05.029.
5
MRT letter: Nanoscopy of protein colocalization in living cells by STED and GSDIM.MRT 信件：通过 STED 和 GSDIM 对活细胞中蛋白质共定位的纳米镜检。
Microsc Res Tech. 2012 Jan;75(1):1-6. doi: 10.1002/jemt.21026. Epub 2011 Jun 15.
6
Stimulated emission depletion nanoscopy of living cells using SNAP-tag fusion proteins.利用 SNAP 标签融合蛋白的活细胞受激发射损耗纳米显微镜技术。
Biophys J. 2010 Jan 6;98(1):158-63. doi: 10.1016/j.bpj.2009.09.053.
7
Low-power compact continuous-wave stimulated emission depletion microscopy.低功率紧凑型连续波受激发射损耗显微镜。
J Biophotonics. 2023 Feb;16(2):e202200233. doi: 10.1002/jbio.202200233. Epub 2022 Sep 13.
8
Diffraction-unlimited all-optical imaging and writing with a photochromic GFP.利用光致变色 GFP 实现无衍射极限的全光学成象和写入。
Nature. 2011 Sep 11;478(7368):204-8. doi: 10.1038/nature10497.
9
Strong signal increase in STED fluorescence microscopy by imaging regions of subdiffraction extent.通过对亚衍射范围区域进行成像，受激发射损耗（STED）荧光显微镜中的信号显著增强。
Proc Natl Acad Sci U S A. 2017 Feb 28;114(9):2125-2130. doi: 10.1073/pnas.1621495114. Epub 2017 Feb 13.
10
An Efficient Green Fluorescent Protein for Stimulated Emission Depletion Super-Resolution Microscopy.一种高效的绿色荧光蛋白，用于受激发射损耗超分辨率显微镜。
Int J Mol Sci. 2022 Feb 24;23(5):2482. doi: 10.3390/ijms23052482.
引用本文的文献
1
Expansion microscopy reveals neural circuit organization in genetic animal models.扩展显微镜技术揭示了基因动物模型中的神经回路组织。
Neurophotonics. 2025 Jan;12(1):010601. doi: 10.1117/1.NPh.12.1.010601. Epub 2024 Dec 20.
2
A pipeline for STED super-resolution imaging and Imaris analysis of nanoscale synapse organization in mouse cortical brain slices.用于 STED 超分辨率成像和 Imaris 分析小鼠皮质脑片纳米级突触组织的流水线。
STAR Protoc. 2023 Dec 15;4(4):102707. doi: 10.1016/j.xpro.2023.102707. Epub 2023 Nov 9.
3
Visualizing and quantifying molecular and cellular processes in Caenorhabditis elegans using light microscopy.使用光学显微镜观察和量化秀丽隐杆线虫中的分子和细胞过程。
Genetics. 2022 Jul 30;221(4). doi: 10.1093/genetics/iyac068.
4
ExCel: Super-Resolution Imaging of C. elegans with Expansion Microscopy.ExCel：利用扩展显微镜对秀丽隐杆线虫进行超分辨率成像。
Methods Mol Biol. 2022;2468:141-203. doi: 10.1007/978-1-0716-2181-3_9.
5
An Efficient Green Fluorescent Protein for Stimulated Emission Depletion Super-Resolution Microscopy.一种高效的绿色荧光蛋白，用于受激发射损耗超分辨率显微镜。
Int J Mol Sci. 2022 Feb 24;23(5):2482. doi: 10.3390/ijms23052482.
6
3D super-resolution deep-tissue imaging in living mice.活体小鼠的三维超分辨率深层组织成像
Optica. 2021 Mar 25;8(4):442-450. doi: 10.1364/OPTICA.416841. eCollection 2021 Apr 20.
7
Ground state depletion microscopy as a tool for studying microglia-synapse interactions.基底状态耗竭显微镜作为研究小胶质细胞-突触相互作用的工具。
J Neurosci Res. 2021 Jun;99(6):1515-1532. doi: 10.1002/jnr.24819. Epub 2021 Mar 7.
8
A Picture Worth a Thousand Molecules-Integrative Technologies for Mapping Subcellular Molecular Organization and Plasticity in Developing Circuits.一张价值千个分子的图片——用于绘制发育中神经回路亚细胞分子组织和可塑性的整合技术
Front Synaptic Neurosci. 2021 Jan 5;12:615059. doi: 10.3389/fnsyn.2020.615059. eCollection 2020.
9
Influence of nanobody binding on fluorescence emission, mobility, and organization of GFP-tagged proteins.纳米抗体结合对绿色荧光蛋白标记蛋白的荧光发射、迁移率和组织的影响。
iScience. 2020 Dec 4;24(1):101891. doi: 10.1016/j.isci.2020.101891. eCollection 2021 Jan 22.
10
Expansion microscopy of .. 的扩展显微镜
Elife. 2020 May 1;9:e46249. doi: 10.7554/eLife.46249.
本文引用的文献
1
The structure of the nervous system of the nematode Caenorhabditis elegans.秀丽隐杆线虫的神经系统结构。
Philos Trans R Soc Lond B Biol Sci. 1986 Nov 12;314(1165):1-340. doi: 10.1098/rstb.1986.0056.
2
Spectroscopic rationale for efficient stimulated-emission depletion microscopy fluorophores.用于高效受激发射损耗显微镜荧光染料的光谱学原理。
J Am Chem Soc. 2010 Apr 14;132(14):5021-3. doi: 10.1021/ja100079w.
3
Subdiffraction-limit two-photon fluorescence microscopy for GFP-tagged cell imaging.用于 GFP 标记细胞成像的亚衍射极限双光子荧光显微镜。
Biophys J. 2009 Dec 16;97(12):3224-8. doi: 10.1016/j.bpj.2009.09.038.
4
Breaking the diffraction resolution limit by stimulated emission: stimulated-emission-depletion fluorescence microscopy.通过受激发射突破衍射分辨率极限：受激发射损耗荧光显微镜技术
Opt Lett. 1994 Jun 1;19(11):780-2. doi: 10.1364/ol.19.000780.
5
STED microscopy with a MHz pulsed stimulated-Raman-scattering source.配备兆赫兹脉冲受激拉曼散射源的受激发射损耗显微镜
Opt Express. 2009 Aug 31;17(18):15679-84. doi: 10.1364/OE.17.015679.
6
Furrow-like invaginations of the yeast plasma membrane correspond to membrane compartment of Can1.酵母质膜的沟状内陷对应于Can1的膜区室。
J Cell Sci. 2009 Aug 15;122(Pt 16):2887-94. doi: 10.1242/jcs.051227. Epub 2009 Jul 28.
7
Super-resolution video microscopy of live cells by structured illumination.通过结构光照明显微镜对活细胞进行超分辨率视频显微镜观察。
Nat Methods. 2009 May;6(5):339-42. doi: 10.1038/nmeth.1324. Epub 2009 Apr 26.
8
Developmental genetics of the C. elegans pharyngeal neurons NSML and NSMR.秀丽隐杆线虫咽神经元NSML和NSMR的发育遗传学
BMC Dev Biol. 2008 Apr 9;8:38. doi: 10.1186/1471-213X-8-38.
9
Ultra-high resolution imaging by fluorescence photoactivation localization microscopy.通过荧光光激活定位显微镜进行的超高分辨率成像。
Biophys J. 2006 Dec 1;91(11):4258-72. doi: 10.1529/biophysj.106.091116. Epub 2006 Sep 15.
10
Imaging intracellular fluorescent proteins at nanometer resolution.以纳米分辨率成像细胞内荧光蛋白。
Science. 2006 Sep 15;313(5793):1642-5. doi: 10.1126/science.1127344. Epub 2006 Aug 10.
Zotero 插件