Leica tauSTED super resolution microscopy for molecular imaging in fixed and live specimens

项目摘要 超分辨荧光显微镜(SRFM)已成为 科学家们将探索离散的分子接近。目前，凯克中心的系统位于 弗吉尼亚大学只能达到≥120纳米的分辨率；要打破固定细胞或活细胞的障碍， Leica Stellaris 8tauSTED显微镜对固定样品的分辨率可达到~30-50 nm。作为一名 研究和教学机构在竞争激烈的学术环境中，做出 这些先进技术可用于生物医学科学。此应用程序的第二个驱动程序 是否需要取代现有的、不可修复的14年前的徕卡SP5x共焦显微镜 系统，于2007年通过S10计划购买。请求的超分辨率tauSTED 显微系统将满足这两方面的需求，并将有益于基础和临床科学 大学；特别是12名参与的联邦资助的调查员(9名主要用户和3名次要用户) 凯克中心(KCCI)。更高的量子效率HYD(46%)和新的光子探测 效率(PDE)检测器(58%)提供了更好的选择，以较少的激发激发样品 并检测细胞和组织中较弱的光子水平。这台仪器对我们的 在纳米尺度上研究亚细胞结构和细胞动力学的核心研究小组 这对培训其他调查人员和参加秋季课程的学生也是至关重要的 学期显微镜课程对各种生物应用的研究感兴趣，包括 发育、神经科学和细胞生物学。 所要求的系统将安装在国际公认的W.M.凯克中心 细胞成像(KCCI)，它以其先进的显微镜和分子技术而闻名 成像。KCCI是一所大学范围内的成像设施，位于所有参与者步行的距离内 来自文理学院、工程学院和医学院的研究人员。KCCI也得到了认可 它的年度实践培训电影和FRET研讨会，在过去的19年里在华盛顿大学举办 弗吉尼亚。所要求的超分辨率tauSTED显微镜是一项先进的技术，它将是 安置在徕卡SP5x房间(11英尺x14英寸)，并进行了适当的配置，以满足实验 本提案中的用户情况以及KCCI成像核心的其他现有和未来用户。 总而言之，所要求的仪器将为加州大学的研究界提供 弗吉尼亚州和更远的地方拥有目前不可用的成像能力，并将增强 重大的正在进行和未来的研究。

Project Summary Super-Resolution Fluorescence Microscopy (SRFM) has become an indispensable tool for scientists to probe discrete molecular proximities. Currently, systems in the Keck Center at the University of Virginia can only reach resolutions of ≥120 nm; to break that barrier for fixed or live cells, Leica STELLARIS 8 tauSTED microscopy can achieve ~30-50 nm resolution for fixed specimens. As a research and teaching institution in the highly competitive academic environment, it is essential to make these advanced technologies available for the biomedical sciences. A second driver for this application is the need to replace the existing and non-repairable 14-year-old Leica SP5x confocal microscopy system, purchased through the S10 program in 2007. The requested super-resolution tauSTED microscopy system will answer both needs and will benefit the basic and clinical sciences of this university; in particular, the 12 participating federally funded investigators (9 major and 3 minor users) of the Keck Center (KCCI). The higher quantum efficiency HyD (46%) and the new photon detection efficiency (PDE) detector (58%) provides a better option to excite the sample with less excitation intensity and detecting weaker photon levels in cells and tissues. This instrument is essential for our core investigator group to investigate the subcellular architecture and cell dynamics at nanoscale dimensions and will also be critical for training other investigators and students attending the fall semester microscopy course interested in the study of various biological applications including developmental, neuroscience and cell biology. The requested system will be housed in the internationally recognized W.M. Keck Center for Cellular Imaging (KCCI), which is particularly known for its advanced microscopy and molecular imaging. KCCI is a university-wide imaging facility, located within walking distance of all participating investigators from the School of Arts & Sciences, Engineering, and Medicine. KCCI is also recognized for its annual hands-on training FLIM & FRET workshop, held for the past 19 years at the University of Virginia. The requested super resolution tauSTED microscopy is an advanced technology, which will be housed in the Leica SP5x room (11'x14') and configured appropriately to meet the experimental conditions of users in this proposal and additional existing and future users of the KCCI imaging core. In summary, the requested instrument will provide the research community at the University of Virginia and beyond with a level of imaging capability that is not currently available and will enhance significantly ongoing and future research.