Core--Microscopy and image analysis

所有提议的项目都需要常规和荧光显微镜，图像分析和延时电影摄影。因此，为了有效地完成我们数据的显微分析，我们需要一个显微核心设备。该设施将配备一名经验丰富的高级研究技术人员（在项目获得资助后招聘），并将协助样品制备、实验执行、图像收集、数据分析以及共聚焦和其他显微图像的编辑。项目负责人同意，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，每个博士后都接受细胞分析设施经理Edward Monosov博士的变形图像分析系统和共聚焦显微镜的使用培训。该培训为项目成员提供操作系统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，也不提供个人独立设置和操作共聚焦显微镜和图像分析系统的深度知识。为了克服这些问题，项目PI想把这些操作交给有经验的显微镜学家/计算机分析员。该人员将接受研究所细胞分析设备经理的培训，并与经理进行沟通，以便为该项目进行共聚焦和图像分析。操作核心将被安置在伯纳姆研究所惠誉大楼（4号楼）一楼一个300平方英尺的房间里，毗邻三个项目实验室。这个房间目前容纳了该计划的405M Axiovert和附带的图像分析系统，以及伯纳姆研究所的第二台共聚焦显微镜，蔡司LSM 410。计划在这个房间里容纳核心项目B的工作人员。一名在显微镜和计算机方面具有丰富经验的核心工作人员将在资助时被招募。工作人员将向Ranscht博士报告，他将担任核心主任（6%的努力）。员工的培训将由研究所细胞分析共享设施的经理Edward Monosov博士提供。伯纳姆研究所的培训时间收费为每小时40美元，共聚焦显微镜的无人监督时间收费为每小时20美元，以抵消MRC1024-MP共聚焦仪器的服务合同和维护费用。1)蔡司405M倒置显微镜及变形图像分析系统。该项目广泛使用倒置蔡司405M显微镜，该显微镜是在1989年获得的现有项目资助的核心。这是一个优秀的显微镜，配备光学相衬，微分干涉对比，暗场，明场和荧光在红色，绿色和紫外线光谱。多年来，该仪器不断更新，配备了新的过滤装置和物镜。所附的外设包括用于培养的加热台和适合于注射培养细胞的埃彭多夫显微注射系统。我们使用Optronics ZVS-47E增强CCD摄像机、Sony Trinitron彩色视频监视器（模拟）和Sony LVR 3000N模拟延时激光光盘录像机扩展了系统的延时电影拍摄能力。

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).