Fluorescence Enhanced Photothermal Infrared Spectroscopy (FE-PTIR) - breakthrough for simultaneous fluorescence microscopy and sub-micron IR spectroscopy

荧光增强光热红外光谱 (FE-PTIR) - 同步荧光显微镜和亚微米红外光谱的突破

• 批准号: 10543927
• 负责人: Craig Prater
• 金额: $ 86.54万
• 依托单位: PHOTOTHERMAL SPECTROSCOPY CORP.
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-04-02 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10543927
• 关键词: Alzheimer' s Disease; Amyloid; Antimicrobial Resistance; Antimicrobial susceptibility; Area; Biochemical; Biocompatible Materials; Biological; Biological Sciences; Biology; Biomedical Engineering; Biomedical Research; Cells; Chemical Structure; Chemicals; Classification; Computer software; Confocal Microscopy; Data; Data Analyses; Dependence; Detection; Development; Disease Resistance; Fluorescence; Fluorescence Microscopy; Glucose; Goals; Heating; Image; In Situ; Individual; Infrared Rays; Institutes; Isotopes; Label; Lasers; Light; Lipids; Malignant Neoplasms; Maps; Measurement; Measures; Metabolism; Microscope; Molecular; Molecular Analysis; Molecular Structure; Multimodal Imaging; Neurodegenerative Disorders; Neurons; Optical Instrument; Pharmaceutical Preparations; Pharmacotherapy; Phase; Photobleaching; Physiologic pulse; Plant Roots; Positioning Attribute; Protein Analysis; Proteins; Publications; Research; Resolution; Sampling; Small Business Innovation Research Grant; Source; Specimen; Spectrum Analysis; Structure; Techniques; Technology; Temperature; Testing; Tissues; United States National Institutes of Health; Visible Radiation; absorption; antimicrobial; biological research; biomedical imaging; brain tissue; cancer cell; commercialization; data acquisition; design; engineering design; fluorescence imaging; fluorophore; glucose metabolism; improved; infrared microscopy; infrared spectroscopy; innovation; insight; interest; microbial; microscopic imaging; multimodality; novel; optical imaging; protein folding; protein misfolding; prototype; response; spectroscopic imaging; submicron; tau Proteins; vibration

1 This Phase II project aims to develop and commercialize Fluorescence Enhanced Photothermal Infrared (FE-PTIR) imaging 2 and spectroscopy. The proposed FE-PTIR will use fluorescence microscopy to map the distribution of fluorescently labeled 3 regions of cells and tissue and then provide chemical structural analysis of the labeled regions using photothermal infrared 4 spectroscopy. Fluorescence microscopy is a cornerstone technique in biological research, allowing sensitive mapping of 5 specifically targeted biomolecules within cells and tissue, but it does not provide information about their molecular 6 structure. Infrared (IR) spectroscopy can provide rich analysis of molecular structure and has been used in life sciences 7 research to study tissue classification, drug/tissue interaction, neurodegenerative diseases, cancer and other areas. 8 Conventional IR spectroscopy, however, has a fundamental spatial resolution limit (i.e. roughly how small an object it can 9 analyze) of around 10 micrometers, similar to the size of an average biological cell. Thus conventional IR spectroscopy has 10 been extremely limited for many biomedical applications where the structures of interest are smaller than a cell. 11 The FE-PTIR technique illuminates a fluorescently labeled sample with UV/visible light which results in fluorescent 12 emission from fluorescently tagged molecules in the sample. A tunable infrared laser source also illuminates the sample, 13 causing localized heating in the sample if the IR laser is tuned to a wavelength that excites molecular vibrations in the 14 sample. Using a camera or other sensitive photodetector is used to record the fluorescent emission from different regions 15 of the sample generates a map of the distribution of fluorescently labeled biomolecules. A key innovation of this proposal 16 was the recognition that common fluorophores have an emission efficiency that is highly temperature dependent. Thus 17 when the sample is also irradiated with infrared light at wavelengths corresponding to molecular vibrations, localized 18 heating from IR absorption by the sample causes a significant change in the fluorescent emission. Recording the emission 19 change as a function of sample position or IR wavelength produces IR absorption images and IR absorption spectra, 20 respectively. Phase I research demonstrated the following key advances: (1) ability of FE-PTIR to map of target 21 biomolecules with fluorescence and analyze the molecular structural of the target molecules; (2) achieve submicron 22 spatial resolution for both fluorescence imaging and infrared spectroscopy; (3) demonstrated a 100X improvement in 23 measurement sensitivity; (4) application of FE-PTIR to study of protein misfolding relevant to neurodegenerative disease 24 research; (5) demonstrated FE-PTIR on individual bacterial and live cancer cells with subcellular resolution. This project is 25 well aligned with NIH goals as it incorporates several key thrusts of the National Institute of Biomedical Imaging and 26 Bioengineering, including optical imaging and spectroscopy, IR imaging, confocal microscopy, and multimodal imaging. FE- 27 PTIR will be extremely useful for example in analyzing the molecular structure/folding/aggregation of fluorescence- 28 localized proteins. Protein misfolding/aggregation is a root cause of many neurodegenerative diseases (e.g. Alzheimer’s). 29 Completion of this Phase II project will lead to the commercialization of a new multimodal microscope that will offer 30 profound benefits for biomedical research including neurodegenerative diseases and antimicrobial resistance research.

1该第二阶段项目旨在开发荧光增强光热红外(FE-PTIR)成像并将其商业化 2和光谱学。拟议的FE-PTIR将使用荧光显微镜来绘制荧光标记的分布 3个细胞和组织的区域，然后利用光热红外提供标记区域的化学结构分析 4光谱学。荧光显微镜是生物学研究的一项基石技术，可以灵敏地绘制出 5专门针对细胞和组织中的生物分子，但它没有提供关于它们的分子的信息 6结构。红外光谱可以提供丰富的分子结构分析，并已在生命科学中得到应用 7研究组织分类、药物/组织相互作用、神经退行性疾病、癌症等领域。 然而，传统的红外光谱有一个基本的空间分辨率限制(即，大致可以有多小的对象 分析)大约10微米，类似于一个普通生物细胞的大小。因此，传统的红外光谱具有 10对于感兴趣的结构比细胞更小的许多生物医学应用来说是极其有限的。 11 FE-PTIR技术用UV/可见光照射带有荧光标记的样品，从而产生荧光 12样品中荧光标记的分子的发射。可调谐红外激光光源也照亮样品， 13如果红外激光被调谐到激发分子振动的波长，则在样品中引起局部加热 14个样本。使用照相机或其他灵敏的光电探测器来记录来自不同地区的荧光发射 15%的样本生成荧光标记生物分子的分布图。这项提议的一项关键创新 16是认识到普通荧光团的排放效率与温度高度相关。因此， 17当样品还被波长与分子振动相对应的红外光照射时，局域化 18样品从红外吸收加热会导致荧光发射的显著变化。记录排放情况 作为样品位置或IR波长的函数的变化产生IR吸收图像和IR吸收光谱， 分别为20个。第一阶段研究展示了以下关键进展：(1)FE-PTIR对靶标的映射能力 21个生物分子的荧光分析和靶分子的分子结构分析；(2)实现亚微米级 22荧光成像和红外光谱的空间分辨率；(3)显示了100倍的 23测量灵敏度；(4)FE-PTIR在神经退行性疾病相关蛋白质错误折叠研究中的应用 24项研究；(5)展示了FE-PTIR对单个细菌和活癌细胞的亚细胞分辨率。这个项目是 25与NIH的目标很好地结合在一起，因为它结合了国家生物医学成像研究所和 26生物工程，包括光学成像和光谱学、红外成像、共聚焦显微镜和多模式成像。铁- 27PTIR将非常有用，例如在分析荧光的分子结构/折叠/聚集- 28个定位蛋白。蛋白质错误折叠/聚集是许多神经退行性疾病(如阿尔茨海默氏症)的根本原因。 29这一第二阶段项目的完成将导致一种新的多模显微镜的商业化，该显微镜将提供 30为包括神经退行性疾病和抗菌素耐药性研究在内的生物医学研究带来深远的好处。