Purchase of a FEI Titan Krios for 3-D EM

描述(由申请人提供)：佛罗里达州立大学(FSU)的生物科学成像资源(BSIR)是佛罗里达州和东南部低温电子显微镜和3-D电子显微镜的焦点。没有其他大学的设施具有类似的重点或设施。这项申请需要资金来更换和升级该设施的中心部件，即带有Titan-Krios的FEI CM 300-FEG电子显微镜，以及具有增强的低温EM和电子断层扫描能力的机器人电子显微镜。我们的CM 300-FEG显微镜已有15年的历史，无论是在停机时间方面，还是在我们实施该领域其他设备和软件开发的能力方面，都显示出了它的年龄。该仪器缺乏成像滤光片，而成像滤光片对目前涉及厚样品的工作至关重要，而且对于电子显微镜的新发展，如相位板和像差校正器的改造，将变得越来越困难。申请的Titan-Krios将支持BSIR的主要用户Kenneth Taylor博士、Kenneth Roux博士和Thomas Roberts博士、新教员Scott Stagg博士以及最近搬到俄勒冈健康与科学大学的长期合作者Michael Chapman博士的实验室来访者的研究。描述了三个小项目，它们将受益于获得现代电子显微镜。泰坦-克里奥斯还将能够支持远程用户，使其有可能向大学以外的用户方便地提供其广泛的功能，并成为东南地区真正的研究资源。使用电子断层扫描和单粒子方法研究薄的、玻璃体的、冷冻的水合物悬浮液是FSU的一个巨大且不断增长的低温EM研究重点，但获得足够的数据，无论是倾斜序列还是预测，目前是许多项目的瓶颈。Titan-Krios的机器人能力以及运行最先进的自动化EM软件(如Leginon)的能力将缓解这一瓶颈。电子断层扫描是三个申请小组的主要重点，而其他两个小组正在开始探索可能性。Titan-Krios的双轴断层成像能力将提高从冷冻水化和快速冷冻、冷冻替代、塑料嵌入的生物标本获得的重建吞吐量和质量。泰坦-克里奥斯将支持美国国立卫生研究院资助的以下研究：(1)肌源性肌肉中的力产生的基本机制，它与心脏肌肉功能有关；(2)平滑肌肌球蛋白调节，它与高血压的健康相关；(3)参与细胞迁移的结构，它与癌症有关；(4)HIV和SIV病毒的结构，这是理解艾滋病的基础；(5)腺相关病毒作为基因治疗载体的发展和(6)NIH对COPII囊泡结构的相关研究，这些囊泡与货物运输缺陷有关，包括乳糜粒滞留病和颅豆状缝发育不良。

DESCRIPTION (provided by applicant): The Biological Science Imaging Resource (BSIR) at Florida State University (FSU) is the focal point for cryoelectron microscopy and 3-D electron microscopy in the state of Florida and the Southeast. No other University facility has a comparable emphasis or facilities. This application requests funds to replace and upgrade the center piece of this facility, an FEI CM 300-FEG electron microscope with a Titan-Krios, a robotic electron microscope with enhanced capabilities for cryoEM and electron tomography. Our CM 300-FEG microscope is 15 years old and is showing its age both in terms of down time and our ability to implement instrumentation and software developments produced by others in the field. The instrument lacks an imaging filter which is crucial for work that now involves thick specimens, and will become increasing problematic for retrofit of new developments in electron microscopy such as phase plates and aberration correctors. The requested Titan-Krios will support the research of the major BSIR users, Drs. Kenneth Taylor, Kenneth Roux and Thomas Roberts, new faculty member Dr. Scott Stagg and visitors from the lab of a long standing collaborator, Dr. Michael Chapman, who recently moved to Oregon Health & Science University. Three minor projects are described that will benefit from access to a modern electron microscope. The Titan-Krios will also be able to support remote users, making it possible to offer its extensive capabilities conveniently to users outside of the university and become a true research resource for the Southeast. The study of thin, vitreous, frozen hydrated suspensions, both using electron tomography and single particle methods is a large and growing cryoEM research emphasis at FSU, but obtaining sufficient data, either as tilt series or projections is currently a bottleneck in a number of projects. The robotic capabilities of the Titan-Krios as well as the ability to run state of the art automated EM software, such as Leginon, will alleviate this bottleneck. Electron tomography is a major emphasis of three applicant groups while the other two are beginning to explore the possibilities. The dual axis tomography capabilities of the Titan-Krios will improve both throughput and quality of reconstructions obtained from both frozen hydrated as well as rapidly frozen, freeze substituted, plastic embedded biological specimens. The Titan-Krios will support NIH funded research in (1) the fundamental mechanism of force generation in a myogenic muscle, which has health relevence to cardiac muscle function; (2) smooth muscle myosin regulation, which has health relevance in hypertension; (3) structures involved in cell migration, which has health relevence in cancer; (4) the structure of HIV and SIV viruses which are fundamental to understanding AIDS; (5) the development of Adeno Associated Virus as a gene therapy vector and (6) NIH related research into the structure of COPII vesicles, which are implicated in defects of cargo trafficking including chylomicron retention disease and cranio-lenticulo-sutural displasia.