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Automation of quantitative DIC microscopy for 3D live-cell imaging using programm

使用程序实现 3D 活细胞成像定量 DIC 显微镜自动化

基本信息

批准号：
8060603
负责人：
Carol Cogswell
金额：
$ 34.4万
依托单位：
BOULDER NONLINEAR SYSTEMS, INC.
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-04-15 至 2012-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8060603
关键词：
Automation Biological Biomedical Research Cells Cellular biology Chemicals Colorado Communities Complex Coupling Development Devices Drosophila genus Dyes Embryo Evaluation Future Image Imaging Techniques In Vitro Investigation Laboratories Length Life Light Marketing Measurement Measures Mechanics Methods Microscope Microscopy Morphologic artifacts Nomarski Interference Contrast Microscopy Onset of illness Optics Performance Phase Positioning Attribute Process Property Refractive Indices Research Resolution Rotation Solutions Speed Staging Staining method Stains System Technology Testing Thick Time Universities Variant absorption active method cellular development cellular imaging cost density design digital imaging fluorescence imaging image processing innovation insight interest liquid crystal meetings millisecond novel optical imaging public health relevance research study success

项目摘要

DESCRIPTION (provided by applicant): The primary objective proposed is to utilize Boulder Nonlinear System's (BNS) spatial light modulator (SLM) technology to automate the Cogswell group's recently-developed quantitative DIC microscope (Q-DIC) and make it accessible to biologists in the marketplace. Q-DIC is a full-field (non-scanning) phase imaging technique which provides optical path length measurements indicative of either thickness, density, or chemical variations. The present limitations of the University of Colorado (CU) lab Q-DIC system can be overcome by adapting BNS spatial light modulators for the microscope in such a way that it will be possible to dynamically vary the three requisite DIC optical design parameters (i.e. phase bias, shear distance and shear direction). A SLM in this context is an electrically driven, anisotropic liquid crystal device with the ability to dynamically change the complex phase of an incident optical wavefront. Precise control of DIC parameter variability using SLM technology will enable automated acquisition of Q-DIC images for non-invasive, real-time quantitative phase (optical path length) imaging. This new capability will allow evaluation of the advantages of Q-DIC for high-resolution biological imaging and will also strengthen the utility of DIC comparisons to fluorescence imaging. Its ability to produce contrast in live-cell images without introducing dyes (which potentially interfere with cellular dynamics) will be of special interest to the cellular biology community. After first proving the feasibility of automating DIC image acquisition with existing SLM technology, BNS proposes to design and fabricate two new and distinctly different SLM configurations in order to compare and contrast their performance and determine which (if either) is best-suited for further development in Phase II. A marketable SLM design must meet the technical challenges of retro-fitting to existing commercial microscopes without limiting their multimode functionality or their high-resolution optical quality. These new configurations will be tested and evaluated through in vitro imaging studies of developing Drosophila embryos, provided by our collaborating biologist (Dr. T. Su). The success of this Phase I research will be measured by the ability to demonstrate accurate quantification of a known cellular development process through a live-cell Q-DIC imaging experiment. In addition to automating the CU lab Q-DIC microscope, this proposal includes initial investigation into whether the new microscope-compatible SLM configurations can also be used more generally for enhancing optical microscope design and performance in the future. Directly coupling new methods of active optical wavefront manipulation to novel methods for digital image processing has the potential to inspire pioneering methods of optimized optical and digital imaging processing. Examples include extending the depth of field of high resolution objectives, active aberration correction, and super-resolution. Such innovations could aid understanding of basic biological problems and biomedical research. PUBLIC HEALTH RELEVANCE: Quantitative differential interference contrast (Q-DIC) microscopy is a full-field-of-view (non-scanning) phase imaging technique which extracts optical path length measurements indicative of either thickness, density, or chemical variations from contrast in images of non-absorbing (or partially absorbing) objects without introducing dyes (which potentially interfere with cellular dynamics). Automated Q-DIC microscopy will enable 3D live-cell Q-DIC imaging and the non-invasive study of dynamic properties during cell development which may reveal new insights into the processes through which the onset of disease takes place.

描述（由申请人提供）：提出的主要目标是利用博尔德非线性系统（BNS）的空间光调制器（SLM）技术来自动化Cogswell集团最近开发的定量DIC显微镜（Q-DIC），并使其在市场上的生物学家可以使用。Q-DIC是一种全场（非扫描）相位成像技术，可提供指示厚度、密度或化学变化的光路长度测量。科罗拉多大学（CU）实验室Q-DIC系统目前的局限性可以通过为显微镜调整BNS空间光调制器来克服，这样就可以动态改变三个必要的DIC光学设计参数（即相位偏差，剪切距离和剪切方向）。在这种情况下，SLM是一种电驱动的各向异性液晶器件，具有动态改变入射光波前的复杂相位的能力。使用SLM技术精确控制DIC参数的可变性，将能够自动获取Q-DIC图像，实现无创、实时定量相位（光路长度）成像。这种新能力将允许评估Q-DIC在高分辨率生物成像方面的优势，并将加强DIC与荧光成像的比较。它在不引入染料（可能干扰细胞动力学）的情况下产生活细胞图像对比度的能力将引起细胞生物学社区的特别兴趣。在首次证明了现有SLM技术自动化DIC图像采集的可行性后，BNS提出设计和制造两种截然不同的新型SLM配置，以比较和对比它们的性能，并确定哪种（如果有的话）最适合第二阶段的进一步开发。一个有市场的SLM设计必须在不限制其多模功能或高分辨率光学质量的情况下，满足现有商用显微镜的改装技术挑战。这些新的配置将通过发育中的果蝇胚胎的体外成像研究进行测试和评估，由我们的合作生物学家（T. Su博士）提供。通过活细胞Q-DIC成像实验，验证已知细胞发育过程的准确定量，将衡量第一阶段研究的成功与否。除了自动化CU实验室的Q-DIC显微镜外，该提案还包括初步研究新的显微镜兼容SLM配置是否也可以在未来更广泛地用于提高光学显微镜的设计和性能。将有源光波前处理的新方法与数字图像处理的新方法直接耦合，有可能激发出优化光学和数字成像处理的开创性方法。例如扩展高分辨率物镜的景深、主动像差校正和超分辨率。这些创新有助于理解基本的生物学问题和生物医学研究。