Leica AOBS SP2 confocal for the MCB/DBS Imaging Facility

描述(由申请人提供)：我们建议购买一台徕卡TCS SP2 AOBS激光扫描共焦显微镜，配备特殊的405 nm二极管和594 nm橙色HeNe激光器，安装在我们现有的光学显微镜成像设备中。在2002年，77名不同的研究人员使用了这一设施，其中56人使用了我们已有五年历史的徕卡SP1 UV共焦。我们用户的增长和日益成熟暴露了现有显微镜的两个重大缺陷。首先是无法执行关键实验，包括1)光漂白后的荧光恢复(FRAP)，2)确定的感兴趣区域中的荧光团的光激活，以及3)以比每1-2秒一个双成分图像更快的速率对多个成分进行时间分辨成像。其次，即使是中等亮度的样品，我们现有的SP1共焦也缺乏敏感度。这使得许多实验更适合在我们的去卷积显微镜上进行共焦，尽管有光学缺陷，只是为了获得足够的灵敏度。解决所有这些问题的一个直接方法是用当代设计取代我们现有的共焦。 在当代的共焦中，徕卡TCS SP2 AOBS和蔡司LSM 510 Meta最适合满足我们最复杂的用户的需求，同时仍然允许多用户设施。我们之所以选择徕卡TCS SP2 AOBS，是因为1)徕卡可以满足我们的实验需求，并具有卓越的灵敏度，在徕卡和蔡司的演示中得到了证明；2)卓越的徕卡“无滤光片”光学设计；3)我们庞大的用户群体的用户界面的连续性，使得不需要进行大量的再培训；4)我们与徕卡现场服务工程师的密切关系的连续性；以及5)我们现有徕卡SP1的价值交易。 NIH资助的主要用户将是Jodi M.Nunnari(《线粒体裂变的调节》)、Jonathon M.Scholey(《基于微管的运动在有丝分裂和细胞内运输中的作用》)和Carol A.Erickson(《活胚胎中神经脊细胞迁移的直接观察》)。这些用户将占仪器时间的大约50%，其余的将提供给我们的大型成像社区。这项使用将继续在一名博士和一名MS科学家的指导下进行，并根据现有部门成像委员会的现行政策进行管理。其中包括一封信，信中表示大力支持维持和继续运作协调中心。

DESCRIPTION (provided by applicant): We propose to purchase a Leica TCS SP2 AOBS laser scanning confocal microscope equipped with special 405 nm diode and 594 nm orange HeNe lasers to be housed in our existing light microscopy Imaging Facility. This facility was used by 77 different researchers in calendar year 2002, including 56 using our fiveyear- old Leica SP1 UV confocal. The growth and increasing sophistication of our users have revealed two significant shortcomings of the existing microscopes. First is the inability to perform key experiments, including 1) fluorescence recovery after photobleaching (FRAP), 2) photo-activation of fluorophores in defined regions of interest, and 3) time-resolved imaging of more than one component at rates faster than one two-component image every 1-2 seconds. Second is a debilitating lack of sensitivity from our existing SP1 confocal for even moderately bright samples. This leads many experiments better suited for a confocal to be done on our deconvolution microscope, despite the optical flaws, simply to get adequate sensitivity. A straightforward way of addressing all of these concerns is the replacement of our existing confocal with one of contemporary design. Of contemporary confocals, the Leica TCS SP2 AOBS and Zeiss LSM 510 META are best suited to meet the needs of our most sophisticated users while still allowing a multi-user facility. We have selected the Leica TCS SP2 AOBS because 1) the Leica can meet our experimental needs and has superior sensitivity as evidenced during demos by Leica and Zeiss; 2) the superior Leica "filter-free" optical design; 3) the continuity of user interface for our large user group, making extensive retraining unnecessary; 4) continuity of our strong relationship with our Leica field service engineer, and 5) the trade in value of our existing Leica SP1. Major NIH-funded users will be Jodi M. Nunnari ("Regulation of Mitochondrial Fission"), Jonathon M. Scholey ("Roles of Microtubule-Based Motility in Mitosis and Intracellular Transport"), and Carol A. Erickson ("Direct Observation of Neural Crest Cell Emigration in a Living Embryo"). These users will account for approximately 50% of the instrument time, with the balance being offered to our large imaging community. This use will continue to be under the direction of one PhD and one MS scientist and administered under the current policies of an existing departmental Imaging Committee. A letter showing strong institutional support for maintenance and continued operation of the confocal is included.