Development of New Automated Polarized Light Microscope

新型自动偏光显微镜的开发

• 批准号: 6915169
• 负责人: RUDOLF OLDENBOURG
• 金额: $ 49.35万
• 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 2009-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6915169
• 关键词: Diptera; bioengineering /biomedical engineering; bioimaging /biomedical imaging; biomedical equipment development; birefringences; cell cycle; centromere; chromosome aberrations; chromosome movement; cytoskeleton; dichroism; fluorescence polarization; liquid crystal; meiosis; microscopy; microtubules; mitotic spindle apparatus; optical polarization; sperm

DESCRIPTION (provided by applicant): We are building and applying an ultra-sensitive and high-resolution polarized light microscope on liquid based crystal technology (the LC-PolScope). We use the unique instrumentation to reveal and decipher the functional role of the dynamic architecture of the cytoskeleton, with a focus on the mitotic apparatus during cell division. The proposed instrumentation continues the development of the LC-PolScope that has greatly enhanced the utility of polarized light for live cell imaging. During the last grant period, we removed an important limitation by adding an aperture scanning device to the microscope optical train. Unlike any other polarizing microscopes, including the LCPolScope, the new Scanned Aperture PolScope can reveal the birefringence of objects that are inclined to the plane of focus. Therefore, this technique can finally reveal birefringent objects, such as mitotic spindles, that have any orientation, and it is not limited to objects that either by chance or by special manipulation are oriented in the plane of focus. Building on the proof of concept, we now propose to refine the aperture scanning technique by modifying the design of the aperture scanner and adding a second, similar device in the imaging path of the microscope. These changes and additions to the optical set-up will make it possible (a) to measure birefringence more accurately and sensitively, and (b) measure dichroism in specimens that exhibit polarization-dependent absorption. Furthermore, with the addition of highly discriminating interference filters to the imaging path, the instrument will be capable of revealing fluorescence polarization. Birefringence, dichroism, and fluorescence polarization are important indicators of anisotropy in local molecular order. Structural anisotropy often proves to be important in establishing biological function, such as the alignment of lipid molecules in forming membranes or the linear arrangement of spindle microtubules for the directed segregation of chromosomes during cell division. By taking advantage of the new and improved capabilities of the Scanned Aperture PolScope, we will investigate mechanisms of chromosome segregation during meiosis in insect spermatocytes. Spermatocytes of Sciara and crane flies provide the opportunity to compare the results from normal chromosome segregation with results obtained with chromosomes that show" either natural (monopolar spindle in Sciara and late segregation of sex chromosomes in crane fly) or induced (cold treatment or drug exposure) abnormalities in their segregation behavior.

描述（由申请人提供）： 我们正在建立和应用基于液晶技术的超灵敏度和高分辨率偏振光显微镜（LC-PolScope）。我们使用独特的仪器来揭示和破译细胞骨架的动态结构的功能作用，重点是细胞分裂过程中的有丝分裂装置。 拟议的仪器继续发展的LC-PolScope，大大提高了实用的偏振光活细胞成像。在上一个资助期间，我们通过在显微镜光学系统上增加孔径扫描装置消除了一个重要的限制。与包括LCPolScope在内的任何其他偏光显微镜不同，新的扫描孔径偏光显微镜可以显示倾斜于焦平面的物体的双折射。因此，这种技术最终可以揭示具有任何取向的双折射物体，例如有丝分裂纺锤体，并且不限于偶然或通过特殊操作在焦平面中取向的物体。 在概念验证的基础上，我们现在建议通过修改孔径扫描仪的设计并在显微镜的成像路径中添加第二个类似的设备来改进孔径扫描技术。这些变化和增加的光学设置将有可能（a）更准确和灵敏地测量双折射，和（B）测量样品中的二向色性，表现出偏振相关的吸收。此外，通过在成像路径中添加高分辨干涉滤光片，该仪器将能够显示荧光偏振。 双折射、二向色性和荧光偏振是局部分子有序各向异性的重要指标。结构各向异性通常被证明在建立生物学功能中是重要的，例如形成膜的脂质分子的排列或在细胞分裂期间染色体定向分离的纺锤体微管的线性排列。通过利用扫描孔径PolScope的新的和改进的功能，我们将研究昆虫精母细胞减数分裂过程中染色体分离的机制。Sciara和起重机蝇的精母细胞提供了一个机会，可以将正常染色体分离的结果与显示”自然（Sciara的单极纺锤体和起重机蝇的性染色体晚期分离）或诱导（冷处理或药物暴露）分离行为异常“的染色体分离结果进行比较。