Core--Microscopy and image analysis

描述所有拟议的项目都需要常规和荧光显微镜，图像分析和延时摄影。因此，为了有效地完成对我们数据的显微分析，我们要求显微镜核心设施。该设施将配备一名经验丰富的高级研究技术人员（在项目获得资助时招聘），并将在样品制备、实验执行、图像收集、数据分析以及共聚焦和其他显微图像编辑方面提供协助。项目PI一致认为，该设施对于有效实现拟议项目的目标至关重要。在我们目前的设置中，细胞分析设施经理Edward Monosov博士对个人博士后进行了使用Metamorph图像分析系统和共聚焦显微镜的培训。该培训为计划成员提供操作系统以解决特定问题的技能。然而，从本质上讲，这种培训既不提供程序内的连续性，也不提供个人独立分离和操作共聚焦显微镜和图像分析系统用于许多应用的知识深度。为了克服这些问题，计划PI希望将这些操作交给经验丰富的显微镜学家/计算机分析师。该人员将接受该研究所细胞分析设施经理的培训并与其沟通，以便为该计划进行共聚焦和图像分析。核心将位于伯纳姆研究所惠誉大楼（4号楼）一楼一个300平方英尺的房间内，毗邻三个计划实验室。这个房间目前容纳该计划的405 M Axiovert和附加的图像分析系统，以及伯纳姆研究所的第二个共聚焦显微镜，蔡司LSM 410。计划是在这个房间里容纳项目核心B的工作人员。当这笔赠款获得资助时，将招募一名在显微镜和计算方面具有丰富经验的核心工作人员。工作人员将向Ranscht博士报告，Ranscht博士将担任核心主任（6%的努力）。工作人员的培训将由该研究所细胞分析共享设施的经理Edward Monosov博士提供。伯纳姆研究所对培训时间收取40美元/小时的费用，对共聚焦显微镜上的无人监督时间收取20美元/小时的费用，以抵消服务合同和维护MRC 1024-MP共聚焦仪器的费用。设备1）Zeiss 405 M倒置显微镜和变形图像分析系统。该计划项目广泛使用倒置蔡司405 M显微镜，该显微镜是在第1年（1989年）获得的现有计划资助的核心。这是一个优秀的显微镜，配备了光学相衬，微分干涉差，暗场，亮场和荧光的红色，绿色和紫外光谱。该仪器多年来一直在更新，增加了新的过滤器和物镜。附加的外围设备包括用于培养的加热台和适于注射培养细胞的Eppendorff显微注射系统。我们已经扩展了该系统的能力，使用Optronics ZVS-47 E增强CCD摄像机，索尼特丽珑彩色视频监视器（模拟）和延时激光视盘记录器（索尼LVR 3000 N，模拟）进行延时电影摄影。

Description All of the proposed projects require conventional and fluorescence microscopy, image analysis, and time lapse cinematography. Thus, to efficiently accomplish the microscopic analysis of our data, we request a microscopy Core Facility. This Facility will be staffed with an experienced senior research technician (to be recruited when the program is funded), and will provide assistance with sample preparation, execution of the experiments, image collection, data analysis and editing of confocal and other microscopic images. The Program PI's concurred that this facility is essential to efficiently achieve the goals of the proposed program. In our current set-up, individual postdocs are trained in the use of the Metamorph Image Analysis system and the confocal microscope by the Manager of the Cell Analysis Facility, Dr. Edward Monosov. This training provides members of the program with skills to operate the systems to address a specific question. By nature, however, this training provides neither continuity within the program, nor depth of knowledge to an individual to independently sept-up and operate the Confocal Microscope and Image Analysis Systems for numerous applications. In order to overcome these problems, the Program PI's want to place these operations in the hands of an experienced microscopist/computer analyst. This person will be trained by and interface with the manager of the Institute's Cell Analysis Facility in order to conduct confocal and image analyses for the Program. Operation The core will be house in a 300 square foot room on the first floor on the Burnham Institute's Fitch Building (Building 4), adjacent to three of the Program Laboratories. This room currently houses the Program's 405M Axiovert and the attached image analysis system, and the Burnham Institute's second Confocal microscope, a Zeiss LSM 410. Plans are to accommodate the staff of Program Project Core B in this room. A core staff person with extensive experience in microscopy and computing will be recruited when this grant is funded. The staff will report to Dr. Ranscht, who will serve as the core director (6% effort). Training of the staff will be provided by Dr. Edward Monosov, the manager of the Institute's Cell Analysis Shared Facility The Burnham Institute charges for training time $40/hour, and for unsupervised time on the confocal microscope $20 per hour to offset costs for service contracts and maintenance of the MRC1024-MP confocal instrument. Equipment 1) Zeiss 405M inverted Microscope and Metamorph Image Analysis System. The Program Project extensively uses an inverted Zeiss 405M Microscope that was acquired for the Core of the existing Program Grant in year 1 (1989). This is an excellent microscope that is equipped with optics for phase contrast, differential interference contrast, darkfield, bright field and fluorescence in the red, green and ultra violet spectra. The instrument has been updated over the years with new filter sets and objectives. Attached peripherals included a heated stage for the cultures and an Eppendorff microinjection system suited for the injection of cultured cell. We have extended the capability of the system to do lapse time cinematography using an Optronics ZVS-47E intensified CCD video camera, a Sony Trinitron Color Video Monitor (analog) and a time lapse laser videodisk recorder (Sony LVR 3000N, analog).