Massively parallel interferometric confocal microscopy using degenerate lasers

使用简并激光器的大规模并行干涉共焦显微镜

• 批准号: 8770650
• 负责人: Hui Cao
• 金额: $ 20.99万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-01 至 2016-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8770650
• 关键词: Biological; Biology; Biomedical Engineering; Cilia; Clinical; Complex; Confocal Microscopy; Detection; Development; Diagnosis; Diagnostic tests; Disease; Doppler Effect; Embryo; Endoscopy; Epidermis; Epithelial; Exhibits; Fiber; Frequencies; Functional disorder; Future; Goals; Grant; Image; Individual; Interferometry; Lasers; Lead; Light; Lighting; Lung diseases; Medical Imaging; Medicine; Methods; Microscope; Microscopy; Motion; Mucous body substance; Optical Coherence Tomography; Optics; Phase; Physiologic pulse; Physiology; Public Health; Pump; Rana; Research; Research Project Grants; Resolution; Respiratory System; Sampling; Scanning; Signal Transduction; Skin; Source; Speed; Structure; Surface; System; Testing; Tissues; Velocimetries; Work; Xenopus; base; bioimaging; design; fluid flow; imaging modality; innovation; new technology; novel; novel diagnostics; operation; particle; public health relevance; respiratory; response; vector; virtual

DESCRIPTION (provided by applicant): Confocal microscopy uses geometric principles to generate cross-sectional images of scattering biological tissue. Traditional reflectance confocal microscopes use serial pixel acquisition, thereby limiting acquisition speeds. Parallelization of pixel acquisition would dramatically increase imaging speeds and enable the study of new kinds of physiology. For example, massively parallelized reflectance confocal microscopy would be invaluable in the emerging field of cilia-driven mucus physiology in the respiratory system. Indeed, in terms of hypothesis-driven ciliary biology, there is a critical methodological gap in high-speed (0.1 to 1 kHz frame rates) imaging methods at the ~1 um scale. Traditional confocal microscopy uses physical pinholes in sample illumination and in photo detection to generate cross-sectional images by straightforward geometric principles. While the use of physical pinholes can be partially parallelized, it is impossible to have complete, scan-free parallelizatio using physical pinholes. However, parallel interferometric confocal microscopy allows an entire field-of-view to be imaged without scanning. In interferometric confocal microscopy, virtual interferometric pinholes are generated, and mutually incoherent spatial modes act as independent and parallelizable confocal imaging channels. However, the lack of sources with low spatial coherence (many independent spatial modes) and high brightness per spatial mode is a critical barrier to massively parallel interferometric confocal microscopy. Traditional lasers exhibit high spatial coherence, while low spatial coherence sources such as thermal sources and light-emitting diodes (LEDs) do not provide the necessary brightness. Recently, we have shown that degenerate Nd: YAG lasers can support as many as ~105 mutually incoherent lasing modes (independent imaging channels). Therefore, using a specifically-designed degenerate Nd: YAG laser, we will build a massively parallel interferometric confocal imaging system with a 100 Hz frame rate for imaging in the ~1 um resolution regime. We expect the principles of operation to allow future scaling into the >kHz frame rate regime. We will use frequency-doubled 532 nm light for interferometric reflectance confocal imaging in ~104 (100 x 100) parallel, independent imaging channels. ~104 imaging channels is chosen as a design specification because several commonly-used imaging fiber bundles used in endoscopy have ~104 imaging cores. As an initial demonstration, we will image ciliary physiology in Xenopus (frog) embryos. Like respiratory epithelial surfaces, the epidermis (skin) of Xenopus embryos is ciliated and generates directional fluid flow. We will image the motion of individual cilia as well quantify cilia-driven fluid flow using interferometry-enabled Doppler flow imaging. Our design-driven research will support the future development of novel diagnostics in respiratory ciliary physiology. Our development of a degenerate laser for confocal microscopy supports future research in degenerate lasers for other kinds of medical imaging (e.g. optical coherence tomography, HiLo structured illumination, holographic microscopy).

描述（由申请人提供）：共聚焦显微镜使用几何原理生成散射生物组织的横截面图像。传统的反射共聚焦显微镜使用串行像素采集，从而限制了采集速度。像素采集的并行化将大大提高成像速度，并使新的生理学研究成为可能。例如，大规模平行反射共聚焦显微镜在呼吸系统中纤毛驱动的粘液生理学的新兴领域将是无价的。事实上，就假设驱动的纤毛生物学而言，在~ 1um尺度上的高速（0.1至1khz帧率）成像方法存在关键的方法差距。传统的共聚焦显微镜使用物理针孔在样品照明和在光检测产生横截面图像由简单的几何原理。虽然使用物理针孔可以实现部分并行化，但使用物理针孔不可能实现完全的、无扫描的并行化。然而，平行干涉共聚焦显微镜允许整个视场成像而无需扫描。在干涉共聚焦显微镜中，产生了虚拟的干涉针孔，相互非相干的空间模式作为独立的、可并行的共聚焦成像通道。然而，缺乏具有低空间相干性（许多独立的空间模式）和每个空间模式高亮度的光源是大规模平行干涉共聚焦显微镜的关键障碍。传统的激光