Core--Microscopy and image analysis

所有拟议的项目都需要常规显微镜和荧光显微镜、图像分析和时间推移电影摄影。因此，为了有效地完成我们的数据的微观分析，我们需要一个显微镜核心设备。该设施将配备一名经验丰富的高级研究技术员(将在项目获得资金时招聘)，并将在样品准备、实验执行、图像收集、数据分析以及共焦图像和其他显微图像的编辑方面提供协助。方案国际一致认为，这一设施对于有效实现拟议方案的目标至关重要。在我们目前的设置中，个别博士后由细胞分析设施经理Edward Monosov博士培训使用变形图像分析系统和共焦显微镜。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，这种培训既不能在项目中提供连续性，也不能为个人提供独立操作共焦显微镜和图像分析系统以应用于多种应用的知识深度。为了克服这些问题，程序PI希望将这些操作交给有经验的显微镜专家/计算机分析师。该人员将接受研究所细胞分析设施经理的培训，并与其接触，以便为该计划进行共焦和图像分析。核心操作将在伯纳姆研究所惠誉大楼(4号楼)一楼300平方英尺的房间内进行，毗邻三个项目实验室。这个房间目前存放着该计划的405M Axiovert和所附的图像分析系统，以及伯纳姆研究所的第二台共焦显微镜--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，将招聘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。工作人员将向兰施特博士汇报工作，兰施特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。Burnham Institute收取的培训时间为每小时40美元，使用共焦显微镜的无人监督时间为每小时20美元，以抵消服务合同和MRC1024-MP共焦仪器的维护成本。设备1)蔡司405M倒置显微镜和变形器图像分析系统。该计划项目广泛使用了一台倒置的蔡司405M显微镜，该显微镜是在第一年(1989)为现有计划拨款的核心而购买的。这是一台出色的显微镜，配备了相位对比度、差分干涉对比度、暗场、亮场以及红、绿和紫外线光谱中的荧光光学元件。多年来，该工具更新了新的过滤器设置和目标。所连接的外围设备包括用于培养的加热平台和适合于注射培养细胞的埃彭多夫微量注射系统。我们已经扩展了系统的功能，可以使用Optronics ZVS-47E增强型CCD摄像机、Sony Trinitron彩色视频监视器(模拟)和延时激光录像机(Sony LVR 3000N，模拟)进行延时电影拍摄。

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).