Zeiss Laser TIRF 3 microscope and live cell imaging system

蔡司激光 TIRF 3 显微镜和活细胞成像系统

• 批准号: 7794499
• 负责人: Joseph E Mazurkiewicz
• 金额: $ 35.2万
• 依托单位: ALBANY MEDICAL COLLEGE
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-04-22 至 2011-04-21
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7794499
• 关键词: Actins; Area; Binding; Biosensor; CDC42 gene; Cell membrane; Cells; Centrosome; Communities; Cytoskeleton; Endocytosis; Event; Fluorescence; Fluorescence Resonance Energy Transfer; Focal Adhesions; Glass; Human; Integrins; Interphase; Lasers; Lead; Life; Ligands; Membrane; Methodology; Microscope; Microtubules; Mitosis; Optics; Phagocytosis; Phagosomes; Production; Proteins; Public Health Schools; Regulation; Research; Research Personnel; Resolution; Resources; Signal Transduction; Signaling Molecule; Site; Smooth Muscle Myocytes; Surface; System; Technology; Total Internal Reflection Fluorescent; Tumor Cell Invasion; Vesicle; adapter protein; base; cell assembly; cell motility; cellular imaging; desensitization; extracellular; fluorescence imaging; fluorophore; instrument; macrophage; medical schools; member; molecular imaging; receptor; success

DESCRIPTION (provided by applicant): This is a request for a Zeiss Laser TIRF 3 microscope and live cell imaging system that will permit a group of 5 researchers at the Albany Medical College and 1 at the SUNY School of Public Health to selectively image molecular events that occur within or very close to the plasma membrane of cells as they lie close to a glass coverslip surface. In TIRF microscopy, a very thin, sub-resolution optical volume about 100 nm in depth is illuminated by the evanescent wave and only fluorescent molecules located within this space are excited, while fluorophores located outside this space do not contribute to the background or the fluorescence signal, resulting in the production of high-contrast, high-resolution fluorescence images. The members of the user group will apply this capability in several diverse areas of study: understanding how integrins regulate centrosome function and microtubule assembly during interphase and mitosis; the regulation of the assembly and disassembly of focal contact between cells and molecules in the extracellular substrate by following the dynamic interaction of fluorescent-protein components in live cells during smooth muscle cell migration and tumor cell invasion; use of the FRET based biosensor, Raichu, to assess the distribution of Rac, CDC42 and RhoA in migrating cells; the assembly of the phagocytic cup as vesicles bringing new membrane to the assembly site, approach and fuse into phagosomes; the interaction of signaling molecules and the actin cytoskeleton during phagocytosis by macrophages as detected with FRET-based biosensors; the interaction of adapter proteins with the human FSHR that leads to stimulation of several signaling cascades and receptor desensitization via endocytosis following binding of its ligand FSH use of the FRET based biosensor, Raichu, to assess the distribution of Rac, CDC42 and RhoA in migrating cells; the optimization of TIRF-FRET methodology. Acquisition of this instrument will dramatically enhance the research endeavors of members of the user group that are in progress. Currently they have no access to a TIRF microscope. The presence of the TIRF microscope will become a central resource in the institutional research community and lead to initiation of new projects as success of the user group using this technology becomes widely known.

描述（由申请人提供）：这是对Zeiss Laser TIRF 3显微镜和活细胞成像系统的申请，该系统将允许奥尔巴尼医学院的5名研究人员和SUNY公共卫生学院的1名研究人员选择性地对细胞质膜内或非常接近细胞质膜的分子事件进行成像，因为细胞靠近盖玻片表面。在TIRF显微镜中，一个非常薄的，亚分辨率光学体积约100 nm的深度是由倏逝波照射，只有位于该空间内的荧光分子被激发，而位于该空间外的荧光团不有助于背景或荧光信号，从而产生高对比度，高分辨率的荧光图像。用户组的成员将在几个不同的研究领域应用这种能力：了解整合素如何调节中心体功能和微管组装在间期和有丝分裂;组装和拆卸的细胞和分子之间的细胞外基质的焦点接触的调节，通过以下的动态相互作用的荧光蛋白组分在活细胞平滑肌细胞迁移和肿瘤细胞侵袭;使用基于FRET的生物传感器Raichu来评估Rac、CDC 42和RhoA在迁移细胞中的分布;吞噬杯作为囊泡的组装，将新膜带到组装位点，接近并融合成吞噬体;信号分子和肌动蛋白细胞骨架在巨噬细胞吞噬期间的相互作用，如使用基于FRET的生物传感器检测的;衔接蛋白与人FSHR的相互作用，其导致在其配体FSH结合后通过内吞作用刺激几个信号级联和受体脱敏，使用基于FRET的生物传感器Raichu评估Rac、CDC 42和RhoA在迁移细胞中的分布; TIRF-FRET方法的优化。获得这一仪器将大大加强正在进行的用户组成员的研究努力。目前，他们还无法使用TIRF显微镜。TIRF显微镜的出现将成为机构研究界的核心资源，并导致新项目的启动，因为使用该技术的用户组的成功变得广为人知。