The National Center for In-situ Tomographic Ultramicroscopy (NCITU)

摘要 我们建议建立国家原位断层扫描超微中心(NCITU)，总部设在 西蒙斯电子显微镜中心(SEMC)，这是纽约结构生物学中心的一部分 (NYSBC)，总部设在纽约。NCITU将致力于开发和提供对这两个标准的访问 和先进的原位冷冻样品制备方法，并通过一个 旨在为各种技能水平和职业目标服务的CryoET交叉培训计划。 NCITU的主要目标将是为用户提供访问和服务，以便为直接 大分子联合和组织的可视化，特别是在天然细胞中。一位非常合格的 技术团队将提供直接动手服务、指导和协助，以确保样品和网格 以最高的质量和尽可能短的时间准备好。已经建立了良好的交叉培训 将扩大和扩大方案，以提供有关冷冻样品准备的各个方面的培训 输油管道。NCITU的业务将建立在SEMC/NYSBC已经建立的做法基础上，并受益于 整个充满活力的环境以及已经到位的训练有素的工作人员的专业知识。 NCITU将为断层扫描的现场样品制备提供全方位的服务，包括：培养 生物样品，柱式冷冻和高压冷冻，低温光学显微镜(低温显微镜)，低温 超微切割、聚焦离子束研磨(FIB-研磨)(包括抬出)和初始CryoTEM筛查 将标本提交给网络中心，以便收集高分辨率的低温冷凝器数据。我们已经这么做了 为其中许多活动建立了工作流程，并有专注于扩展的积极研究项目 自动化和简化整个工作流程的功能。我们将继续走在新的前沿 我们将把新的方法、技术和工作流程纳入到标准中 NCITU的操作程序只要这些都是稳定和稳健的。 虽然NCITU的目标并非与生俱来的创新性，但我们的意图是将该标准定为第一个 世界一流的、可在全国范围内使用的冷冻样品制备设施，以提供访问和培训，即 免费和公平地提供给美国所有的研究人员，并继续努力改进方法和使 最高质量和最有价值的科学。

Summary We propose to establish the National Center for In-situ Tomographic Ultramicroscopy (NCITU) to be based at the Simons Electron Microscopy Center (SEMC), which is a part of the New York Structural Biology Center (NYSBC), based in New York, NY. NCITU will be dedicated to developing and providing access to both standard and advanced methods for in-situ cryoET specimen preparation, and to disseminating this expertise through a cryoET cross-training program designed to serve a wide variety of skill levels and career goals. The primary goal of NCITU will be to provide user access and service towards preparing specimen for direct visualization of macromolecular associations and organization, particularly in native cells. A highly qualified technical team will provide direct hands-on service, guidance, and assistance to ensure that samples and grids are prepared with the highest quality and in the shortest time possible. An already well established cross-training program will be expanded and extended to provide training in every aspect of the cryoET specimen preparation pipeline. The operations at NCITU will build on already established practices at SEMC/NYSBC and benefit from the overall vibrant environment as well as the expertise of the highly trained staff already in place. NCITU will provide a full range of services for in-situ sample preparation for tomography including: culturing biological samples, plunge freezing and high pressure freezing, cryo-light microscopy (cryoLM), cryo- ultramicrotomy, focused ion beam milling (FIB-milling) including lift-out, and initial cryoTEM screening prior to submission of specimens to the network Hub for high-resolution cryoET data collection. We have already established workflows for many of these activities and have active research projects focused on expanding capabilities for automating and streamlining the entire workflow. We will continue to stay at the forefront of new developments in this area and we will incorporate new methods, technologies and workflows into the standard NCITU operating procedures as soon as these are stable and robust. While the goals of NCITU are not innately innovative, it is our intention to set the standard as one of the first world class, nationally accessible cryoET sample preparation facilities, to provide access and training that is freely and fairly available to all researchers in the US, and to continually strive to improve methods and enable science of the highest quality and value.