JEOL JEM-1400 Tomographic Transmission EM with High-Resolution Digital Cameras

JEOL JEM-1400 配备高分辨率数码相机的断层扫描透射电子显微镜

• 批准号: 7838715
• 负责人: JOHN E RASH
• 金额: $ 80.43万
• 依托单位: COLORADO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-01 至 2012-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7838715
• 关键词: Actins; Archives; Binding; Boxing; Colorado; Electron Microscope; Electrons; Ensure; Exanthema; Excision; Film; Freeze Fracturing; Freezing; Frequencies; Funding; Gap Junctions; Image; Label; Laboratories; Liquid substance; Mechanics; Medicine; Membrane; Molecular; Monitor; National Center for Research Resources; Neurodegenerative Disorders; Neuronal Plasticity; Neurons; Nitrogen; Operative Surgical Procedures; Peer Review; Persons; Photography; Potassium Channel; Productivity; Publications; Publishing; Research Personnel; Resolution; Scanning; Schedule; Specimen; Staging; Surface; Surveys; Synapses; Time; Universities; Viral Proteins; Virion; Virus Receptors; Work; Wyoming; base; cofilin; college; digital; digital imaging; experience; neurotransmission; receptor; reconstruction; repaired; research study; retinal rods; stereoscopic; trafficking; transmission process

DESCRIPTION (provided by applicant): Five established and three new NIH-funded investigators from three departments at Colorado State University, plus one new investigator from the University of Wyoming, and one established investigator via a subcontract from Albert Einstein College of Medicine, request $804,260 in NCRR funds to purchase a JEOL JEM-1400 120 kV transmission electron microscope (TEM) equipped with a liquid nitrogen-cooled 700 tomographic specimen tilt holder, a fast-scan digital camera, and a high-resolution slow-scan digital camera. The new TEM will replace a 19-yr-old JEOL JEM-2000 EX-II 200 kV "workhorse" TEM that lacks a digital camera, has no tomographic capability, and has no automated stage drives and no automated photographic capability. Its "cut- film" camera has made >67,000 negatives (>67,000 mechanical cycles), requiring us to purchase three new camera boxes because the worn mechanical advance frequently jammed, irrevocably damaging three. The JEM-2000 has had increasing frequency of repairs ("down time" of 20-38 working days in each of the last five years. The requested bottom-mount high-resolution digital cameras, in combination with automated tilting and automated photography, are essential for tomographic analysis (up to 70 images for each reconstruction) and will greatly facilitate conventional photography. Digital images will be stored centrally for access online or offline without possibility of altering or deleting the archived originals. The video-rate 11MB camera will facilitate focusing and rapid tracking of image detail on the monitor during group analysis, and will be the primary camera for obtaining routine survey micrographs. Single-image capture of hundreds of virus particles using the 4K (16MB) high-resolution digital camera (as opposed to 2K or 1K cameras) will greatly reduce beam damage to virus/receptor-protein interactions in frozen thin films by reducing by 4x or 16x the number of exposures needed for tomographic analysis. On-line stereoscopic imaging, often at extreme tilts, is a necessity for interpreting surface topography and the "sidedness" of immunogold-labels in freeze-fracture replicas, rather than waiting until the next day for film to be developed. Removal of this delay between experiments will greatly accelerate work in several laboratories. The high productivity of current "major users" has resulted in >1100 published electron micrographs from the JEM-2000 in >50 peer-reviewed publications. TEM is essential for 6 NIH-funded major users investigating molecular composition of neuronal and glial membranes (Rash), ADF/cofilin-actin rods in neurodegenerative disease (Bamburg), synaptic and non-synaptic mechanisms of neuronal transmission (Reist; Rash), virus/protein molecular interactions (Hafenstein), regulators of K+ channel function and trafficking (Tamkun), and plasticity of neuronal gap junctions (Pereda/Rash). Based on 19 years of experience with the JEM-2000 TEM, a 10%-time administrative and technical person (Ms. Davidson) and a broadly-based Advisory Board will ensure continued efficient scheduling and operation of the upgraded replacement transmission electron microscope.

描述（由申请人提供）：来自科罗拉多州立大学三个系的 5 名固定研究员和 3 名新的 NIH 资助研究员，加上怀俄明大学的一名新研究员，以及通过阿尔伯特爱因斯坦医学院分包合同确定的一名固定研究员，请求 NCRR 资金 804,260 美元购买配备液氮冷却的 JEOL JEM-1400 120 kV 透射电子显微镜 (TEM) 700断层标本倾斜支架、快速扫描数码相机和高分辨率慢速扫描数码相机。新的 TEM 将取代已有 19 年历史的 JEOL JEM-2000 EX-II 200 kV“主力”TEM，该 TEM 缺乏数码相机，没有断层扫描功能，没有自动载物台驱动器，也没有自动摄影功能。它的“剪辑”相机已经制作了超过 67,000 张底片（>67,000 次机械循环），要求我们购买三个新的相机盒，因为磨损的机械推进器经常卡住，不可挽回地损坏三个相机盒。 JEM-2000 的维修频率不断增加（过去五年每年的“停机时间”为 20-38 个工作日。所需的底部安装高分辨率数码相机与自动倾斜和自动摄影相结合，对于断层扫描分析至关重要（每次重建最多 70 张图像），并将极大地方便传统摄影。数字图像将集中存储，以便在线或离线访问，而无需更改或离线 删除存档的原件。视频速率 11MB 相机将有助于在群体分析期间在监视器上聚焦和快速跟踪图像细节，并将成为获取常规调查显微照片的主要相机。使用 4K (16MB) 高分辨率数码相机（相对于 2K 或 1K 相机）捕获数百个病毒颗粒的单图像将大大减少冷冻薄层中病毒/受体-蛋白质相互作用的光束损伤 将断层扫描分析所需的曝光次数减少 4 倍或 16 倍。在线立体成像（通常处于极端倾斜状态）对于解释冷冻断裂复制品中的表面形貌和免疫金标签的“侧面”是必要的，而不是等到第二天才冲洗胶片。消除实验之间的这种延迟将大大加快几个实验室的工作速度。的高生产率 目前的“主要用户”已在超过 50 个同行评审出版物中发表了超过 1100 张来自 JEM-2000 的电子显微照片。 TEM 对于 6 个 NIH 资助的主要用户至关重要，这些用户研究神经元和神经胶质膜的分子组成（皮疹）、神经退行性疾病中的 ADF/丝切蛋白肌动蛋白杆（Bamburg）、神经元传递的突触和非突触机制 (Reist; Rash)、病毒/蛋白质分子相互作用 (Hafenstein)、K+ 通道功能和运输的调节剂 (Tamkun) 以及神经元间隙连接的可塑性 (Pereda/Rash)。基于 19 年 JEM-2000 TEM 的经验，一名占 10% 时间的行政和技术人员（戴维森女士）和一个基础广泛的顾问委员会将确保该项目的持续高效调度和运行。 升级更换透射电子显微镜。