Stanford-SLAC CryoET Specimen Preparation Service Center (SCSC)

斯坦福-SLAC CryoET 标本制备服务中心 (SCSC)

• 批准号: 10264893
• 负责人: Wah Chiu
• 金额: $ 169.6万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-16 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10264893
• 关键词: Academia; Adopted; Advisory Committees; Area; Award; Biocompatible Materials; Biological; Biomedical Research; Cells; Cellular Structures; Collaborations; Communication; Communities; Complex; Cryo-electron tomography; Cryoelectron Microscopy; Data Collection; Educational Curriculum; Educational workshop; Equipment; Evaluation; Fluorescence Microscopy; Freezing; Funding; Goals; Grant; Hydration status; In Situ; Industry; Infrastructure; Institution; Investigation; Ions; Laboratories; Lifting; Macromolecular Complexes; Methods; Microscope; Molecular Structure; Organelles; Preparation; Process; Professional Organizations; Protocols documentation; Request for Applications; Research; Research Personnel; Resolution; Resource Allocation; Resources; Sampling; Scanning Electron Microscopy; Schedule; Scientist; Selection Criteria; Services; Specimen; Subcellular structure; Techniques; Technology; Tissue Sample; Tissues; Titan; Training; Training Activity; United States National Institutes of Health; Work; cryogenics; data management; electron tomography; experience; experimental study; imaging modality; innovative technologies; instrument; instrumentation; interest; member; operation; programs; response; structural biology; symposium; tool; web portal; Academia; Adopted; Advisory Committees; Area; Award; Biocompatible Materials; Biological; Biomedical Research; Cells; Cellular Structures; Collaborations; Communication; Communities; Complex; Cryo-electron tomography; Cryoelectron Microscopy; Data Collection; Educational Curriculum; Educational workshop; Equipment; Evaluation; Fluorescence Microscopy; Freezing; Funding; Goals; Grant; Hydration status; In Situ; Industry; Infrastructure; Institution; Investigation; Ions; Laboratories; Lifting; Macromolecular Complexes; Methods; Microscope; Molecular Structure; Organelles; Preparation; Process; Professional Organizations; Protocols documentation; Request for Applications; Research; Research Personnel; Resolution; Resource Allocation; Resources; Sampling; Scanning Electron Microscopy; Schedule; Scientist; Selection Criteria; Services; Specimen; Subcellular structure; Techniques; Technology; Tissue Sample; Tissues; Titan; Training; Training Activity; United States National Institutes of Health; Work; cryogenics; data management; electron tomography; experience; experimental study; imaging modality; innovative technologies; instrument; instrumentation; interest; member; operation; programs; response; structural biology; symposium; tool; web portal

In response to the NIH RFA-RM-19-009, we propose to establish the Stanford-SLAC CryoET Specimen Preparation Service Center (SCSC) to accomplish four specific aims: (1) establish a platform, which is available to the scientific community at large, to streamline the preparation of samples suitable for downstream cryogenic electron tomography (cryoET) data collection; (2) provide access to advanced cryo-specimen preparation techniques for a wide range of samples: macromolecular complexes, microcrystals of biological materials, cell lysates, organelles, cells and tissues; (3) generate a training curriculum for new users on the preparation of frozen, hydrated biological specimens, with the option of carrying out correlative cryo-fluorescence light microscopy (cryoFLM), cryo-focused ion beam scanning electron microscopy (cryoFIB), and cryoET; and (4) adopt new methods for sample preparation using innovative technologies developed elsewhere. In this proposal, we will leverage our existing facilities and in-house expertise in each of the following imaging modalities and their integration into a correlated sample preparation workflow: cryoFLM, cryoFIB, and cryoET. We will establish all the necessary equipment dedicated to the proposed service and training activities. We will prepare video curriculum materials to train new users in the aforementioned protocols. We will adapt our existing infrastructure to manage project administration and resource allocations. Administrative support is in place to help users with lodging arrangements, as well as laboratory on-boarding and specimen biosafety approvals. We have a well set- up communication infrastructure for remote users to participate in actual experiments, particularly for cryoFLM, cryoFIB, and cryoET. We will provide access to existing Talos Arctica and/or Titan Krios microscopes to evaluate whether the prepared samples are ready for data collection in the associated Hub, which will be set up at another institution with separate NIH support. We will make our existing Stanford-SLAC cryo-specimen preparation equipment available on a limited basis immediately after this proposal is funded. We anticipate serving ~40 users per year once the Center is fully operational in early Year 2 of the award period. We will also offer regular hands- on workshops to train ~10-12 new users per year. We will disseminate our resources to the broad community via a web portal and booths at professional societies’ annual conferences. The number of users will increase in subsequent years as we and our recurring users optimize the use of the Center’s resources. In Years 3-6, we will work closely with collaborators pioneering advanced protocols such as semi-automated and automated cryoFIB and “lift-out” for tissue samples to implement them in the proposed Center. Lastly, we will assemble a Scientific Advisory Committee of experts to guide the practices of our Service Center and the ongoing implementation of cutting-edge technologies.

为了响应NIH RFA-RM-19-009，我们建议建立Stanford-SLAC冷冻标本 准备服务中心（SCSC）实现了四个具体目标：（1）建立一个平台，可用 到整个科学界，以简化适合下游低温的样品的准备 电子断层扫描（冷冻）数据收集； （2）提供对高级冷冻特性准备的访问 广泛样品的技术：大分子复合物，生物材料的微晶，细胞 裂解物，细胞器，细胞和组织； （3）为新用户生成培训课程 冷冻的水合生物学标本，可以选择执行相关的冷冻荧光光 显微镜（Cryoflm），以冷冻关注的离子束扫描电子显微镜（Cryofib）和冷冻; （4） 采用新方法，使用其他地方开发的创新技术进行样品制备。在此提案中， 我们将利用以下每种成像方式及其的现有设施和内部专业知识 集成到相关的样品制备工作流程中：Cryoflm，Cryofib和Cryoet。我们将建立所有 专门用于拟议的服务和培训活动的必要设备。我们将准备视频 课程材料在预测协议中培训新用户。我们将调整现有的基础架构 管理项目管理和资源分配。为帮助用户提供管理支持 住宿安排，以及实验室的登机和标本生物安全批准。我们有一个很好的设置 - 为远程用户提供通信基础架构参与实际实验，特别是对于CryoFLM， 冰冻和冷冻。我们将提供对现有的Talos Arctica和/或Titan Krios显微镜的访问权限 准备好的样品是否准备好在关联中心中进行数据收集，该样本将在另一个设置 机构有单独的NIH支持。我们将做我们现有的斯坦福 - 塞拉克冷冻式制剂 在资助该提案后，立即以有限的方式提供设备。我们预计将为约40个用户提供服务 每年该中心在奖励期的第二年中完全运行。我们还将提供常规的手 - 在研讨会上，每年培训约10-12个新用户。我们将把我们的资源传播给广泛的社区 通过Web门户网站并在专业社会的年度会议上展位。用户的数量将增加 随后的几年，我们和我们的经常性用户优化了中心资源的使用。在3 - 6年中，我们 将与合作者开创性高级协议（例如半自动化和自动化）紧密合作 冰冻纤维和“提出”，以便在拟议的中心中实现纸巾样品。最后，我们将组装一个 科学咨询专家委员会指导我们服务中心的实践和正在进行的实践 实施尖端技术。