A Cell Dynamics Microscope for Live Cell Fluorescence

描述(申请人提供)：本申请的目标是建立一种新的最先进的设备，称为细胞动力学显微镜，用于使用荧光显微镜观察和操纵活细胞内的分子。设想的工作站以蔡司Axiovert 200M倒置显微镜为中心，带有两个主要附件。第一种是Perkin Elmer Ultraview活细胞成像仪，用于成像活细胞中的荧光，光漂白和光毒性最小。第二个附件是来自光电子仪器的数字光圈激光照明系统。这一新仪器允许对视野内的任何子区域、子区域集合或图案进行强激光照明。通过在目标的计算机监视器图像上绘制或导入图案来控制子区域或图案的设置。这种能力允许对光化学技术的光照进行前所未有的控制，包括光漂白后荧光恢复(FRAP)、光漂白中的荧光损失(FLIP)、荧光共振能量转移(FRET)、笼状化合物的光活化、直接光烧蚀、染料敏化光烧蚀(有时称为生色团辅助激光灭活或CALL)和离子成像。主要用户组的成员将在几个领域应用这些技术：有丝分裂中的微管和动粒动力学(Gorbsky)，感光细胞中的蛋白质转位(McGinnis)，胰岛素介导的GLUT4转位中的微管(Olson)，膀胱上皮中的核因子-KB动力学(Saban)，以及初生纤毛和有丝分裂纺锤体中的多囊蛋白-2(Tsiokas)。这些项目还将得到一组额外用户的九个项目的补充。使用这种新仪器进行的分析将补充每个实验室正在进行的分子研究。细胞动力学显微镜将成为研究人员社区的关键资源，他们寻求优雅和用户友好的方法来观察和操纵活细胞内的目标分子。

DESCRIPTION (provided by applicant): The objective of this application is to establish a novel state-of-the-art facility termed the Cell Dynamics Microscope for the observation and manipulation of molecules within living cells using fluorescence microscopy. The envisioned workstation is centered on a Zeiss Axiovert 200M inverted microscope with two major accessories. The first is a Perkin Elmer Ultraview Live Cell Imager for imaging fluorescence in living cells with minimal photobleaching and phototoxicity. The second accessory is a Digital Diaphragm laser illumination system from Photonic Instruments. This new instrument allows intense laser illumination of any subregion, set of subregions or pattern within the field of view. Setting of subregions or patterns is controlled by drawing or importing patterns on a computer monitor image of the target. This ability permits unprecedented control of illumination for photochemical techniques including fluorescence recovery after photobleaching (FRAP), Fluorescence loss in photobleaching (FLIP), fluorescence resonance energy transfer (FRET), photoactivation of caged compounds, direct photoablation, dyesensitized photoablation (sometimes called chromophore-assisted laser inactivation or CALl), and ion imaging. The members of the major user group will apply these techniques in several areas: Microtubule and Kinetochore Dynamics in Mitosis (Gorbsky), Protein Translocation in Photoreceptor Cells (McGinnis), Microtubules in Insulin-mediated Glut4 Translocation (Olson), NF-KB Dynamics in Bladder Epithelium (Saban), and Polycystin-2 in Primary Cilia and Mitotic Spindles (Tsiokas). These projects will be supplemented by nine projects from a group of additional users. The analyses performed with this new instrument will complement the molecular studies ongoing in each laboratory. The Cell Dynamics Microscope will become a pivotal resource for the community of researchers seeking elegant and userfriendly methods to observe and manipulate target molecules within living cells.