DEVELOPMENT OF AUTOMATED POLARIZED LIGHT MICROSCOPE

自动偏光显微镜的研制

• 批准号: 6052377
• 负责人: RUDOLF OLDENBOURG
• 金额: $ 41.28万
• 依托单位: MARINE BIOLOGICAL LABORATORY
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-08-12 至 2002-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6052377
• 关键词: bioimaging /biomedical imaging; biomedical automation; biomedical equipment development; birefringences; cytology; cytoskeleton; fertilization; fluorescence polarization; image enhancement; image processing; light microscopy; liquid crystal; nucleic acid structure; optical polarization; optical rotation; structural biology

Given adequate sensitivity, polarizing microscopes provide the means to measure submicroscopic molecular arrangements in living cells and other specimens. The polarizing microscope can thus provide information which is otherwise only available with more intrusive techniques such as electron microscopy. We have been developing and applying a new polarized light microscope (Pol-Scope) which dramatically enhances the unique capabilities of the traditional polarizing microscope. The Pol- Scope incorporates a universal compensator, made from computer driven liquid crystal devices, to measure the birefringent fine structure for all orientations of the birefringence axis within the plane of focus. We propose to develop a 2nd generation Pol-Scope by improving the design and function of the universal compensator and by adding the capability for measuring the magnitude and orientation of birefringences in all three dimensions, including those structures whose birefringence axis do not lie in the focal plane. To increase sensitivity, we will refine the design of the universal compensator by using sectored liquid crystal plates to rectify polarization errors introduced by high numerical aperture lenses. With the rectified Pol-Scope we expect to be able to measure extremely low levels of birefringence such as exhibited by single actin filaments. Also, the rectified system is needed for the 3-D capability of birefringence measurement. To realize the capability of measuring the complete 3-D birefringence indicatix for each resolvable area in the specimen, we will place a rotatable, asymmetric mask (or its equivalent sectored liquid crystal plate) into the back focal plane of the condenser lens. The asymmetric mask will effectively tilt the light path through the specimen. Thus, data for birefringence components parallel to the optical axis of the microscope will become available. Data will be recorded for several mask positions and image records will be combined using digital processing for a complete 3-D analysis of molecular orientation and birefringent fine structure. To test the accuracy and sensitivity of the 2nd generation Pol- Scope, we will use a variety of man-made and biological model systems, including small birefringent crystals and isolated asters. We will extend our collaborative applications projects on cytoskeletal dynamics in motile cells, DNA tertiary structure in chromosomes, and spindles and zona pellucida of mammalian oocytes and their relation to oocyte infertility.

如果有足够的灵敏度，偏光显微镜提供了测量活细胞和其他标本中亚微观分子排列的手段。 因此，偏光显微镜可以提供信息，而这些信息只能通过电子显微镜等更具侵入性的技术才能获得。我们一直在开发和应用一种新的偏光显微镜（Pol-Scope），它大大增强了传统偏光显微镜的独特功能。 Pol- Scope包含一个通用补偿器，由计算机驱动的液晶器件制成，用于测量焦平面内双折射轴所有方向的双折射精细结构。我们建议通过改进通用补偿器的设计和功能，并通过增加测量所有三维中双折射的大小和方向的能力，包括双折射轴不在焦平面中的结构，来开发第二代Pol-Scope。为了提高灵敏度，我们将改进通用补偿器的设计，通过使用扇形液晶板来校正由高数值孔径透镜引入的偏振误差。 与整流Pol-Scope，我们希望能够测量极低水平的双折射，如单肌动蛋白丝表现出。 此外，校正系统是必要的三维双折射测量能力。为了实现测量样品中每个可分辨区域的完整3-D双折射率的能力，我们将在聚光透镜的后焦平面中放置可旋转的非对称掩模（或其等效的扇形液晶板）。 非对称掩模将有效地倾斜通过样品的光路。 因此，平行于显微镜光轴的双折射分量的数据将变得可用。 数据将被记录为几个掩模位置和图像记录将结合使用数字处理的分子取向和双折射精细结构的完整的3-D分析。为了测试第二代Pol- Scope的准确性和灵敏度，我们将使用各种人造和生物模型系统，包括小的双折射晶体和孤立的星状体。 我们将扩大我们的合作应用项目在运动细胞的细胞骨架动力学，染色体中的DNA三级结构，纺锤体和哺乳动物卵母细胞的透明质酸及其与卵母细胞不育的关系。