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

总结 我们建议建立国家原位层析超显微镜中心（NCITU）， 西蒙斯电子显微镜中心（SEMC）是纽约结构生物学中心的一部分 （NYSBC），总部位于纽约州纽约。NCITU将致力于开发和提供两种标准的接入， 和先进的原位冷冻ET标本制备方法，并通过一个 cryoET交叉培训计划旨在服务于各种技能水平和职业目标。 NCITU的主要目标将是为用户提供访问和服务，以准备标本供直接使用。 大分子缔合和组织的可视化，特别是在天然细胞中。一支高素质 技术小组将提供直接的实践服务、指导和协助，以确保样品和网格 在最短的时间内以最高的质量准备。一个已经建立的交叉训练 该计划将扩大和扩展，以提供冷冻ET标本制备各个方面的培训 渠道. NCITU的运营将建立在SEMC/NYSBC已经建立的实践基础上，并受益于 整个充满活力的环境以及训练有素的工作人员的专业知识已经到位。 NCITU将为断层扫描原位样品制备提供全方位的服务，包括： 生物样品，插入式冷冻和高压冷冻，低温光学显微镜（cryoLM），低温， 超薄切片术、聚焦离子束研磨（FIB研磨）（包括剥离）和冷冻前的初始cryoTEM筛选 向网络中心提交标本，以收集高分辨率cryoET数据。我们已经 为许多这些活动建立了工作流程，并有积极的研究项目，重点是扩大 自动化和简化整个工作流程的能力。我们将继续保持在新的前沿 我们将把新的方法、技术和工作流程纳入标准 NCITU的操作程序，只要这些是稳定和强大的。 虽然NCITU的目标并不是天生的创新，但我们的意图是将该标准设定为最早的标准之一 世界一流的，全国可访问的冷冻ET样品制备设施，提供访问和培训， 免费和公平地提供给美国的所有研究人员，并不断努力改进方法，使 最高质量和价值的科学。

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.