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（申请人提供）：来自科罗拉多州立大学三个系的五名已成立和三名新的NIH资助的研究人员，加上一名来自怀俄明州大学的新研究人员，以及一名来自阿尔伯特爱因斯坦医学院的研究人员，要求804美元，260 NCRR资金用于购买JEOL JEM-1400 120 kV透射电子显微镜（TEM），配备液氮冷却700断层扫描试样倾斜保持器，快速扫描数码相机，和高分辨率慢扫描数码相机。新的TEM将取代19年前的JEOL JEM-2000 EX-II 200 kV“主力”TEM，该TEM没有数码相机，没有层析成像能力，没有自动载物台驱动器，也没有自动摄影能力。它的“切割胶片”相机已经制作了超过67,000张底片（超过67,000个机械循环），我们需要购买三个新的相机盒，因为磨损的机械推进经常卡住，不可避免地损坏了三个。JEM-2000型飞机的修理频率越来越高（在过去五年中，每年的“停机时间”为20至38个工作日）。所要求的底部安装高分辨率数码相机，结合自动倾斜和自动摄影，对于断层分析（每次重建多达70张图像）至关重要，并将大大促进传统摄影。数字图像将集中存储，以便在线或离线访问，而不可能更改或删除存档的原件。视频速率为11 MB的摄像机将有助于在组分析期间在监视器上聚焦和快速跟踪图像细节，并且将是获取常规调查显微照片的主要摄像机。使用4K（16 MB）高分辨率数码相机（而不是2K或1 K相机）捕获数百个病毒颗粒的单个图像将大大减少冷冻薄膜中病毒/受体-蛋白质相互作用的光束损伤，减少4倍或16倍断层扫描分析所需的曝光次数。在线立体成像，往往在极端的倾斜，是一个必要的解释表面形貌和“侧”的免疫金标签在冷冻断裂复制品，而不是等到第二天的电影被开发。消除实验之间的这种延迟将大大加快几个实验室的工作。目前“主要用户”的高生产率已导致在50多份同行评审的出版物中发表了1 100多张JEM-2000电子显微照片。TEM对于6个NIH资助的主要用户研究神经元和神经胶质膜的分子组成（Rash），神经退行性疾病中的ADF/cofilin-actin棒（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）至关重要。基于19年使用JEM-2000 TEM的经验，一名10%时间的行政和技术人员（Davidson女士）和一个基础广泛的咨询委员会将确保持续有效地安排和操作升级后的替代透射电子显微镜。