Orientation independent DIC and polarization microscopy

方向无关的 DIC 和偏振显微镜

• 批准号: 7197108
• 负责人: MICHAEL SHRIBAK
• 金额: $ 32.59万
• 依托单位: MARINE BIOLOGICAL LABORATORY
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-02-01 至 2009-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7197108
• 关键词: Algorithms; Anisotropy; Behavior; Biological; Birefringence; Cells; Cellular Structures; Chromosomes; Computer software; Coupled; Development; Devices; Elements; Event; Fiber; Film; Fluorescence; Generations; Goals; Image; Investigation; Label; Legal patent; Life; Light Microscope; Measurement; Meiosis; Microscope; Microtubules; Mitosis; Molecular; Morphology; Movement; Optics; Organelles; Phase; Polarization Microscopy; Process; Range; Refractive Indices; Research Personnel; Rotation; Scheme; Specimen; Staining method; Stains; System; Techniques; Time; Tissues; cell motility; design; fluorescence imaging; improved; insight; liquid crystal; molecular assembly/self assembly; molecular dynamics; programs; prototype

DESCRIPTION (provided by applicant): This project aims to develop and apply a new, orientation-independent differential interference contrast (OI-DIC) microscope, whose contrast is independent of specimen orientation and is appropriate for investigation of live biological specimens. Extending the preliminary studies using conventional DIG coupled with specimen rotation, by which the theoretical validity of the proposed technique was validated, OI-DIC systems are proposed which require no rotation of either the specimen or the DIG prisms. With the new systems (patent applied), an OI-DIC image should be obtained in 2 seconds or less. Thus microscopes incorporating the new system will allow precise analyses of organellar morphology, motility, as well as dry mass distribution. In addition, a combination of regular DIG and orientation independent polarization (LC-Pol) microscope will be developed. This new system will yield two complementary phase images of thin optical sections of the specimen: distribution of optical path gradients and distribution of birefringence due to structural or internal anisotropy of the cell structure. The DIG and LC-Pol devices combined into one unit will allow rapid switching between the two modes without the need to move any optical components so that both images can be captured nearly simultaneously and without misalignment, yielding a combined image in ca. 0.3 seconds. The possibilities of combining the proposed technique with near-contemporaneous fluorescence images without moving any optical elements will be explored. This combination will give unique views of a specimen, where structural information can be joined with molecularly specific labels. The new devices will be used to study the architectural and molecular dynamics of live biological specimen, with emphasis on events associated with mitosis and meiosis; the OI-DIC providing enhanced chromosome images while the LC-Pol images quantitatively depicting the dynamic assembly-disassembly and distribution of microtubules in the spindle fibers and astral rays. The new microscope should generally allow advanced analyses of organellar movement and sub-organellar anisotropy that reflect the submicroscopic and ongoing molecular events, directly in living cells. RELEVANCE: In order to gain improved insight into the dynamic behavior and molecular events underlying healthy and pathologically impaired tissues, we will develop new optics and processing software for light microscopes that allow speedy capture of informative images without the need to destroy or stain the cells. The new systems will make cell organelles, including their changes and movement more visible, allow measurement of their dry mass, and concurrently reveal changes in molecular assembly and alignment, all non-invasively without perturbing the state of the cells.

描述(申请人提供)：该项目旨在开发和应用一种新型的、与方向无关的差分干涉对比(OI-DIC)显微镜，其对比度与样品方向无关，适合于活体生物标本的研究。扩展了常规DIG与样品旋转相结合的初步研究，验证了所提出技术的理论有效性，提出了不需要旋转样品或DIG棱镜的OI-DIC系统。使用新系统(申请专利)，OI-DIC图像应该在2秒或更短的时间内获得。因此，安装了新系统的显微镜将能够精确分析细胞器的形态、运动性以及干质量分布。此外，还将开发一种规则的Dig和方向无关偏振(LC-Pol)相结合的显微镜。这种新的系统将产生样品的薄光学切片的两个互补的位相图像：光路梯度分布和由于细胞结构的结构或内部各向异性而引起的双折射分布。组合成一个单元的DIG和LC-POL设备将允许在两种模式之间快速切换，而不需要移动任何光学部件，因此可以几乎同时捕获两种图像，并且不会错位，从而在大约0.3秒内产生组合图像。将探索在不移动任何光学元件的情况下将所提出的技术与近同时荧光图像相结合的可能性。这种组合将给出标本的独特视角，其中结构信息可以与分子特定的标记结合在一起。新设备将用于研究活的生物标本的结构和分子动力学，重点是与有丝分裂和减数分裂有关的事件；OI-DIC提供增强的染色体图像，而LC-POL图像定量描述纺锤体纤维和星形射线中微管的动态组装、拆解和分布。新的显微镜一般可以对细胞器的运动和亚细胞器的各向异性进行高级分析，直接在活细胞中反映亚微观和正在进行的分子事件。相关性：为了更好地洞察健康和病理受损组织背后的动态行为和分子事件，我们将为光学显微镜开发新的光学和处理软件，允许快速捕获信息丰富的图像，而不需要破坏或染色细胞。新系统将使细胞器，包括它们的变化和运动更明显，允许测量它们的干质量，并同时揭示分子组装和排列的变化，所有这些都是非侵入性的，不会扰乱细胞的状态。