UltraViewRS Spinning Disk Confocal Microscopy

描述（由申请人提供）：该提案解决了W.M.对额外共聚焦显微镜能力和能力的需求。凯克生物成像设施位于怀特黑德研究所。这个核心设施提供共聚焦培训和服务，20怀特黑德实验室，39从麻省理工学院和4从其他机构共计3732小时的使用之间的1998年和2003年。现有的ZeissLSM 510-Meta基于点扫描技术，因此在图像采集速度方面受到根本限制。在评估了几种竞争性技术，这些技术将解决对额外共聚焦能力的需求以及对快速生物过程成像的能力，我们确定Perkin Elmer的第二代旋转盘系统UltraViewRS是最合适的。该系统将允许用户以超过现有LSM 510 10倍的速率收集时间分辨的2D或3D共焦数据。虽然LSM 510-Meta仍将是该机构的核心技术，特别适用于对厚固定组织样本进行成像，但UltraViewRS提供的补充功能将使更多和更广泛的重要生物学研究得以开展。在项目描述部分，我们描述了已经在7个Whitehead和3个MIT实验室进行的研究，这些实验室需要这些增强的共聚焦能力。这些研究项目包括细胞运动、胚胎发育、微生物发病机理、蛋白质构象疾病、衰老、膜动力学和红细胞生成过程中细胞骨架结构的研究。所提出的系统将允许激发相关的活细胞探针，包括所有主要的GFP变体以及常用的化学探针和固定细胞荧光团。除了提高数据收集的速度外，该系统还通过减少样品光损伤而使用户受益，这对研究胚胎发育的研究人员特别有用。系统的另外两个关键组成部分，一个环境室和一个机动XY载物台，将为该设施提供急需的活细胞能力，并通过自动化多点数据收集制度有效利用该仪器。Whitehead研究所在维护和进一步投资NIH资助的共享仪器方面有着良好的记录，这在W.M.内资源的不断增加的使用中得到了证明。凯克生物成像设施。

DESCRIPTION (provided by applicant): This proposal addresses the need for additional confocal microscopy capacity and capability at the W.M. Keck Biological Imaging Facility located within the Whitehead Institute. This core facility provides confocal training and service to 20 Whitehead labs, 39 from MIT and 4 from other institutions totaling 3732 hours of use between 1998 and 2003. The existing ZeissLSM510-Meta is based on point-scanning technology and is thus fundamentally limited in image acquisition speed. After evaluating several competing technologies that would address the need for additional confocal capacity as well as the ability to image fast biological processes, we determined that the second generation spinning-disk system from Perkin Elmer, the UltraViewRS, was most appropriate. This system will allow users to collect time-resolved 2D or 3D confocal data at over 10x the rate of the existing LSM510. While the LSM510-Meta will remain a core technology within the Facility, especially suited to imaging thick fixed tissue samples, the complementary capability provided by the UltraViewRS will enable an additional and broad range of important biological investigations to be undertaken. In the project descriptions section, we have described studies already underway in 7 Whitehead and 3 MIT laboratories that require these enhanced confocal capabilities. These projects include investigation of cytoskeletal structure during cell motility, embryonic development, microbial pathogenesis, protein conformational diseases, senescence, membrane dynamics, and erythropoiesis. The proposed system will allow excitation of relevant live-cell probes including all major GFP variants as well as commonly-used chemical probes and fixed-cell fluorophores. Apart from the enhanced speed of data collection, the proposed system also benefits users via reduced sample photo-damage, which is especially useful for those investigators studying embryonic development. Two other key system components, an environmental chamber and a motorized XY-stage, will provide much needed live-cell capabilities to the Facility and allow efficient use of the instrument through automated multi-point data collection regimes. The Whitehead Institute has an excellent track record of maintaining and further investing in NIH-funded shared instrumentation as documented in the ever-increasing use of the resources within the W.M. Keck Biological Imaging Facility.