Ultrahigh-resolution and single-molecule stimulated Raman scattering (SRS) microscopy

超高分辨率单分子受激拉曼散射 (SRS) 显微镜

• 批准号: 10377375
• 负责人: Wei Min
• 金额: $ 31.78万
• 依托单位: COLUMBIA UNIV NEW YORK MORNINGSIDE
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-04-01 至 2023-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10377375
• 关键词: Address; Adopted; Algorithms; Alkynes; Amino Acids; Amplifiers; Area; Basic Science; Beds; Biological; Biophotonics; Blinking; Cancer Biology; Carbon; Cells; Chemical Structure; Chemicals; Color; Computational algorithm; Coupling; Crystallization; Detection; Development; Dyes; Fluorescence; Fluorescence Microscopy; Generations; Glucose; Goals; Home; Image; Immunology; Label; Lasers; Lipids; Medical Research; Methods; Microscope; Microscopy; Molecular Target; Neurosciences; Optics; Pharmaceutical Preparations; Property; Pump; Radial; Reporting; Resolution; Sampling; Scheme; Specificity; Speed; System; Techniques; Technology; Testing; Tubulin; base; bioimaging; biological research; biomaterial compatibility; biomedical imaging; complex biological systems; design; detector; diffraction of light; disease diagnostic; imaging platform; improved; instrument; instrumentation; interdisciplinary approach; invention; molecular scale; multiplexed imaging; next generation; novel; precision medicine; second harmonic; single molecule; vibration

Project summary Super-resolution optical microscopy promises to revolutionize biological imaging, as it enables non- invasive interrogation at molecular scale. Indeed, the emergence of super-resolution fluorescence microscopy has quickly impacted the way biologists study cells and subcellular phenomenon. However, super-resolution fluorescence microscopy has fundamental limitations due to the use the fluorescence as contrast mechanism. In particular, it has three major limitations: (1) it cannot reveal chemical composition of the sample; (2) it cannot interrogate small biomolecules due to the relatively bulky fluorescent tags; (3) it cannot image a large number of targets due to the color barrier (only 2~5 fluorescent colors can be practically resolved). The goal of this project is to develop a novel super-resolution imaging platform by exploiting stimulated Raman scattering (SRS) as the contrast mechanism. During the past 10 years since its invention in 2008, SRS microscopy has made widespread impact in biomedical imaging. Being a chemically sensitive method, SRS is well known for its label-free chemical analysis in a quantitative manner. With the recent development of tiny bio-orthogonal tags such as alkynes, SRS has been proven successful in interrogating a wide spectrum of small biomolecules such as lipids, glucose, amino acids, and drugs. Very recently, novel vibrational dye palettes with fine spectral resolution have been reported to achieve super-multiplex electronic pre-resonance (epr) SRS imaging of more than 20 targets simultaneously. Importantly, all these utilizes of SRS microscopy is limited by light diffraction. With SRS being a perfectly complementary contrast mechanism to the prevalent fluorescence, the current proposal aims to develop the necessary methods to bring SRS microscopy to the realm of super resolution. (1) How to improve the resolution for general chemical imaging and small biomolecule imaging; (2) how to break the diffraction limit of the super-multiplex epr-SRS imaging; (3) how to develop the matching vibrational dyes for single molecule SRS. Towards these goals, we had laid out a systematic plan as to how to crystallize this concept into a powerful technology platform. An inter-disciplinary approach has been planned out including instrumentation development, computational imaging, and novel probes synthesis. In Aim 1, we will develop and build new instrumentation. In Aim 2, we will explore new computational algorithm. In Aim 3, we will design next- generation vibrational probes. If successfully implemented, we will establish a transformative platform. The resulting super-resolution chemical imaging would find wide applications in systematically unraveling complex biological systems such as neuroscience, immunology and cancer biology for basic research, disease diagnostics and precision medicine. 1

项目摘要 超分辨率光学显微镜有望彻底改变生物成像，因为它使非生物成像成为可能。 分子级别的侵入性审讯事实上，超分辨率荧光显微镜的出现 迅速影响了生物学家研究细胞和亚细胞现象的方式。 然而，超分辨率荧光显微镜由于使用荧光显微镜而具有根本的局限性。 荧光作为对比机制。特别是，它有三个主要的局限性：（1）它不能揭示化学 （2）由于相对庞大的荧光物质，它不能询问小的生物分子; 标签;（3）由于颜色障碍，它不能对大量目标成像（只能显示2~5种荧光颜色）。 实际解决）。 本项目的目标是利用受激超分辨成像技术，研制一种新型的超分辨成像平台 拉曼散射（SRS）作为对比机制。自2008年发明以来的10年里，SRS 显微术在生物医学成像中产生了广泛的影响。作为一种化学敏感的方法，SRS 以其定量方式的无标记化学分析而闻名。随着最近的发展， 生物正交标签，如炔，SRS已被证明成功地询问了广泛的 小的生物分子，如脂类、葡萄糖、氨基酸和药物。最近，新型振动染料 已经报道了具有精细光谱分辨率的调色板以实现超多路电子预共振 (epr)同时对20多个目标进行SRS成像。重要的是，所有这些利用SRS显微镜是 受到光衍射的限制。 由于SRS是普遍荧光的完美互补对比机制， 目前的建议旨在开发必要的方法，使SRS显微镜的领域， 分辨率(1)如何提高普通化学成像和生物小分子成像的分辨率;（2） 如何突破超多路epr-SRS成像的衍射极限：（3）如何发展匹配 用于单分子SRS的振动染料。 为了实现这些目标，我们制定了一个系统的计划，如何将这一概念具体化为一个 强大的技术平台。一个跨学科的方法已经计划出来，包括仪器 开发、计算成像和新型探针合成。在目标1中，我们将开发和建立新的 仪器仪表在目标2中，我们将探索新的计算算法。在目标3中，我们将设计下一个- 产生振动探针。如果成功实施，我们将建立一个转型平台。的 由此产生的超分辨率化学成像将在系统地揭示复杂的 生物系统，如神经科学，免疫学和癌症生物学的基础研究，疾病 诊断学和精准医学。 1

项目成果

Background-free imaging of chemical bonds by a simple and robust frequency-modulated stimulated Raman scattering microscopy

通过简单而强大的调频受激拉曼散射显微镜对化学键进行无背景成像

• DOI: 10.1364/oe.391016
• 发表时间: 2020
• 期刊: Optics Express
• 影响因子: 3.8
• 作者: Xiong, Hanqing;Qian, Naixin;Zhao, Zhilun;Shi, Lingyan;Miao, Yupeng;Min, Wei
• 通讯作者: Min, Wei