Toward fast and deep imaging of living tissue with cellular resolution

以细胞分辨率对活体组织进行快速、深度成像

• 批准号: 10651713
• 负责人: Nozomi Nishimura
• 金额: $ 62.33万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-07-01 至 2026-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10651713
• 关键词: Address; Adoption; Anatomy; Animal Model; Animals; Biological; Biological Process; Biology; Biomedical Research; Blood capillaries; Brain; Brain region; Cancer Biology; Cardiac; Cell Differentiation process; Cell physiology; Cells; Chronic; Color; Data; Databases; Development; Dyes; Face; Fiber; Fluorescence Microscopy; Frequencies; Gases; Generations; Heart; Hippocampus; Image; Imaging Techniques; Immunology; Injury; Intestines; Kidney; Label; Lasers; Location; Mammary gland; Marrow; Measurement; Measures; Microscope; Microscopy; Mus; Muscle; Neurosciences; Operative Surgical Procedures; Organ; Organ Model; Osteocytes; Penetration; Performance; Perfusion; Photons; Physiologic pulse; Preparation; Process; Proteins; Protocols documentation; Research Activity; Resolution; Signal Transduction; Skeletal Muscle; Skin; Source; Structure; System; Techniques; Testing; Time; Tissue imaging; Tissues; Variant; Vision; Visualization; adaptive optics; biological systems; body system; bone; cell motility; cost; daughter cell; design; experimental study; flexibility; fluorophore; frontier; imaging capabilities; improved; in vivo; instrument; intravital imaging; knowledge base; lymph nodes; mechanical load; neural; new technology; next generation; non-invasive optical imaging; novel; programs; scale up; stem cell fate; timeline; tumor; two photon microscopy; two-photon; waveguide

Abstract An exciting recent development for high spatial resolution deep tissue imaging is long wavelength three- photon fluorescence microscopy (3PM). Since its first demonstration of imaging subcortical structures in the mouse brain, 3PM has driven rapid progress in deep tissue imaging beyond the depth limit of two-photon fluorescence microscopy (2PM). Long-wavelength 3PM is perhaps the most promising new technology for deep imaging within scattering biological tissues, and has potential impacts in a large number of biomedical fields such as neuroscience, immunology, and cancer biology. On the other hand, there are a number of challenges that must be overcome before 3PM can reach its full potential. Because it is a higher-order nonlinear process, three- photon excitation (3PE) is inherently weaker than two-photon excitation (2PE). The weak signal strength of 3PM is particularly problematic for fast imaging of dynamic cellular process. Furthermore, the laser sources for 3PM are not yet optimized for deep tissue penetration, and the complexity and cost of the excitation source is a major barrier for the applications of 3PM in a typical biomedical research lab. Finally, nearly all 3PM applications today are in the brains. Reaching anatomical frontiers is equally possible in other organs with 3PM, but explicit demonstrations of intravital imaging in novel locations are needed to bring deep imaging capability to other biological systems. The research activity of this proposal will directly address the above challenges for in vivo deep tissue 3PM. We will develop a new generation of 3PM that will improve the performance of existing 3PM by two orders of magnitude and enable multi-color deep tissue imaging with a single excitation wavelength. We will demonstrate the unprecedented imaging capabilities with a low-cost, fiber-based laser system, removing a key barrier for the deployment of 3PM in biology labs. Furthermore, by applying our techniques to a wide variety of biological systems, we will create a valuable knowledge base for the applications of 3PM. Our development of the next generation 3PM parallels the development of 2PM, where the concerted development effort in lasers, microscopes, and biological applications in the 1990s made 2PM ubiquitous in biomedical research labs by the early 2000s. Our vision is to make deep, fast 3PM a routine instrument for a wide variety of biomedical applications just as 2PM does in the shallower regions of biological tissues and organs. The successful completion of this program will enable visualization of dynamic process at the sub-cellular level in intact organs and animal models that are completely beyond the reach of any existing imaging techniques.

抽象的 高空间分辨率深层组织成像的令人兴奋的最新发展是长波长3- 光子荧光显微镜（3pm）。自从其首次演示成像中皮质结构以来 小鼠大脑，下午3点已驱动深层组织成像的快速进展，超出了两光子的深度极限 荧光显微镜（2pm）。长波长下午3点可能是最有希望的新技术 在散射生物组织中进行成像，并在许多生物医学领域具有潜在的影响 作为神经科学，免疫学和癌症生物学。另一方面，存在许多挑战 必须在下午3点之前克服其全部潜力。因为它是一个高阶非线性过程，所以 光子激发（3PE）本质上比两光子激发（2PE）弱弱。 3pm的信号强度弱 对于动态细胞过程的快速成像特别有问题。此外，下午3点的激光源 尚未针对深层组织渗透进行优化，激发源的复杂性和成本是主要的 下午3点在典型的生物医学研究实验室中应用的障碍。最后，今天几乎所有下午3点申请 在大脑中。在下午3点的其他器官中，达到解剖边界是同样的，但明确的 需要在新颖位置进行浸润成像的演示，以将深层成像能力带给其他 生物系统。该提案的研究活动将直接解决上述体内挑战 深组织3pm。我们将开发新一代下午3点，以提高现有3pm的性能 通过两个数量级，并启用具有单个激发波长的多色深层组织成像。我们 将通过低成本，基于纤维的激光系统展示前所未有的成像功能 生物学实验室中下午3点部署的关键障碍。此外，通过将我们的技术应用于各种各样 在生物系统中，我们将为下午3点的应用创建一个宝贵的知识库。我们的发展 下一代下午3点与下午2点的开发相似，在这里，激光的共同开发工作， 显微镜和1990年代的生物应用在生物医学研究实验室中无处不在。 2000年代初。我们的愿景是使深度，快速3pm成为常规工具，用于各种生物医学 在生物组织和器官的较浅区域中，施用就像下午2点一样。成功 该程序的完成将使在完整器官的亚细胞级别上可视化动态过程 和动物模型完全无法实现任何现有的成像技术。