Expansion Optical Coherence Microscopy (ExOCM)

扩展光学相干显微镜 (ExOCM)

• 批准号: 10668523
• 负责人: Chao Zhou
• 金额: $ 19.46万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-01 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10668523
• 关键词: 3-Dimensional; Adopted; Adoption; Biomedical Research; Blood Vessels; Cell Communication; Cell Differentiation process; Cells; Color; Communities; Contrast Media; Data; Developmental Biology; Diameter; Dimensions; Dyes; Fluorescence; Growth; Hydrogels; Image; Imaging Device; Label; Light; Microscope; Microscopy; Modeling; Molecular; Molecular Profiling; Optical Coherence Tomography; Optics; Organoids; Process; Proteins; Protocols documentation; Regenerative Medicine; Research; Resolution; Sampling; Scanning; Signal Transduction; Specimen; Speed; Structure; System; Techniques; Technology; Thick; Three-Dimensional Image; Time; Tissue Expansion; Tissue Sample; Tissues; Tumor Angiogenesis; Water; absorption; angiogenesis; antibody conjugate; anticancer research; cell type; detection sensitivity; experience; experimental study; fluorescence microscope; imaging approach; imaging capabilities; imaging platform; imaging system; improved; instrument; interest; meter; microscopic imaging; multiplexed imaging; nanoGold; nanometer resolution; new technology; organ growth; programs; structural imaging; superresolution imaging; technology development; tool; tumor; two photon microscopy; two-photon; ultra high resolution

Project Summary Inspired by recent advances in expansion microscopy (ExM), we propose to develop a new technology called expansion optical coherence microscopy (ExOCM). Using a standard optical coherence tomography (OCT) or a high-resolution OCM system, we will obtain super-resolution, molecular-specific images of expanded tissue samples. Our recent pilot experiment demonstrated that intrinsic scattering signals from expanded samples can be observed using OCT, demonstrating the feasibility of ExOCM based on intrinsic scattering contrast. To further enable molecular-specific imaging, contrast agents, such as gold nanoparticles (GNPs), molecular dyes, and even fluorescence proteins, with different light absorption profiles can be selected for multi-color ExOCM. Compared to standard ExM, ExOCM can image extra thick samples at high speed. This approach also allows the co-registration of three-dimensional (3D) molecular profiles with live tissue microstructures obtained from the same specimen using label-free OCT. Building upon our group’s extensive expertise in developing OCT and OCM technologies and experience with 3D organoid models, we propose to develop, optimize and validate ExOCM technology, and demonstrate the unique attributes of ExOCM for biomedical research. Two specific aims of the proposed program are: Aim 1. To develop and optimize ExOCM technology for multi-color, super- resolution imaging of expanded organoids. We will optimize an ultrahigh-resolution OCM system to achieve ~1.5 µm isotropic image resolution. Antibody conjugated GNPs will be used to provide selective targeting for specific proteins. Using our optimized OCM system and a ~4.5X tissue expansion protocol, we expect to obtain a ~300 nm resolution in both the axial and transverse dimensions. Multi-color and multiplexed imaging will be obtained using spectroscopic OCT analysis to separate signals from GNPs with different absorption and scattering profiles. Aim 2. To validate ExOCM with two-photon microscopy, and to demonstrate the feasibility of the combined OCT and ExOCM imaging approach to characterize 3D structural and molecular profiles and growth dynamics from the same samples. We will use OCT to acquire longitudinal, label-free images of living organoids over a three-week time-course. At predetermined time points, samples will be expanded for ExOCM and two- photon imaging, where tissue structural landmarks will be identified from 3D image stacks for registration. ExOCM imaging data collected during the expansion process will provide first-hand information about 3D expansion uniformity. Complementary 3D structural and molecular profiles and growth dynamics from the same organoids will be obtained, demonstrating the unique advantages of the combined OCT and ExOCM imaging approach. If successful, the integrated OCT and ExOCM imaging platform will prove helpful for applications in developmental biology, cancer research and regenerative medicine. Furthermore, high-throughput characterization of the specimens can be performed using commercially available OCT systems, making ExOCM easy to disseminate and adopt by the broader research community.

项目摘要 受扩展显微镜（EXM）的最新进展的启发，我们建议开发一种新技术，称为 膨胀光学显微镜（EXOCM）。 高分辨率OCM系统，我们将获得扩展组织的超分辨率，分子特异性图像 样本。我们最近的试点实验表明 可以使用OCT观察，这证明了基于内在散射的对比度的可行性 启用分子特异性成像，对比剂，例如金纳米颗粒（GNP），分子染料，以及 甚至可以选择具有不同光吸收曲线的荧光蛋白进行多色EXOCM。 与标准EXM相比，Exocm可以以高速图像额外的厚样品。 三维（3D）分子曲线的共同注册与从TEE获得的活组织微生 使用无标签的OCT的同一标本在我们小组的广泛专业知识上 OCM技术和3D器官模型的经验，我们建议开发，优化和验证 Exocm技术，并展示了生物医学研究的独特属性 支撑计划的目的是：目标1。开发和优化用于多色，超级的Exocm技术 分辨率扩展的器官。我们将优化超高分辨率OCM系统 µM各向同性图像分辨率。 蛋白质使用优化的OCM系统和〜4.5倍的组织扩展方案 将获得轴向和横向尺寸的NM分辨率。 使用光谱OCT分析将单独的信号与具有不同吸收和散射的GNP分开 剖面。 联合OCT和EXOCM成像方法来表征3D结构和分子曲线以及生长 来自同一样品的动力学。 在预定时间点的三周时间里，将扩大样本 光子成像，其中将从3D图像堆栈中识别组织结构地标。 在扩展过程中收集的EXOCM成像数据提供了有关3D的第一手信息 扩展均匀性。 将获得类器官，证明了联合OCT和EXOCM成像的独特优势 方法。 发育生物学，癌症研究和再生医学。 可以再次使用CommerMercial可用OCT执行样品的表征，使ExOCM 易于传播和采用更广泛的研究社区。