Ceramic Electron Microscopy Grids for Cell Culturing and Multiscale Imaging

描述(申请人提供)：需要改进细胞内分子和超分子结构成像的支持技术，以促进细胞生物学研究，国家普通医学科学研究所(具体地说，其细胞生物学和生物物理系)正在寻求这一技术。近年来，利用冷冻电子断层扫描(CRYO-ET)对超分子结构和单分子进行多尺度成像，已被证明是高通量表征细胞动态三维结构的一种独特而宝贵的方法。电子显微镜(EM)栅格被用作支撑被成像的生物和生物分子样品的衬底，是与这种成像方法相关的关键部件。新的EM网格技术降低了样品准备成本和时间，改善了从培养到冷冻的样品生成，并提高了成像质量，将使研究人员能够以更高的吞吐量更有效地探索细胞结构。Synkera提出了一种新型的陶瓷EM栅格，该栅格具有集成的薄支撑膜，与细胞培养、光学显微镜和冷冻高度兼容。网格将促进亚蜂窝架构的高通量、多尺度成像，并提供与最先进产品相比的关键优势。在许多其他EM和培养应用中，网格也有望成为具有竞争力的替代方案。为期6个月的第一阶段项目成功地展示了拟议网格体系结构的可行性，以及它们在培养、冷冻和多尺度成像方面的潜力。第二阶段旨在通过进一步开发制造工艺和陶瓷EM栅格设计，在这一成功的基础上，充分实现功能原型的多尺度成像潜力。至少有四个学术合作伙伴将协助这一发展进程。拟议项目的最终目标是建立一系列完整的陶瓷电磁网格，用于从生物成像到材料表征等广泛的电磁应用。 与公共卫生相关：该项目致力于通过低温电子断层扫描(CRYO-ET)对分子和细胞进行成像。具体地说，目标应用是通过光学显微镜和低温对细胞、超分子和单分子结构进行多尺度成像，以生成亚细胞结构的3D模型。提出了一种新的陶瓷基电子显微镜栅格的开发，以促进这种多尺度成像。拟议的技术将提供比最先进产品更大的能力，并通过简化样品准备过程和产生卓越的成像性能，帮助进一步简化多尺度细胞成像。

DESCRIPTION (provided by applicant): Improved supporting technologies for imaging of molecular and supramolecular structures within cells are needed to facilitate cell biology research, and are sought by the National Institute of General Medical Sciences (specifically, its Division of Cell Biology and Biophysics). Multiscale imaging, using cryo-electron tomography (cryo-ET) on supramolecular structures and single molecules, has proven in recent years to be a unique and invaluable method for highthroughput characterization of the dynamic 3D architecture of cells. Electron microscopy (EM) grids, used as substrates for supporting the biological and biomolecular specimens being imaged, are a critical component associated with this imaging method. New EM grid technology that decreases sample preparation cost and time, improves sample generation from culturing to freezing for cryo-ET, and increases imaging quality will allow researchers to more efficiently explore cellular architecture, at higher throughput. Synkera proposes a novel class of ceramic EM grids that feature an integrated thin support film that is highly compatible with cell culturing, light microscopy and cryo-ET. The grids will facilitate high-throughput, multiscale imaging of sub-cellular architecture and offer key advantages over state-of-the art products. The grids are also expected to be a competitive alternative in many other EM and culturing applications a 6-month Phase I project successfully demonstrated feasibility of the proposed grids architecture, as well as their potential in culturing, cryo-ET and multiscale imaging. Phase II aims to build on this success by further developing fabrication processes and ceramic EM grids designs, to fully realize the potential for multiscale imaging in functional prototypes. At least four academic partners will aide in this development process. The ultimate goal of the proposed project is a complete line of ceramic EM grids for a broad range of EM applications, from bioimaging to materials characterization. PUBLIC HEALTH RELEVANCE: The project addresses imaging of molecules and cells via cryo-electron tomography (cryo-ET). Specifically, the target application is multiscale imaging via optical microscopy and cryo-ET of cellular, supramolecular and single-molecule structures, for generating 3D models of sub-cellular architecture. The development of a novel class of ceramic-based electron microscopy grids that facilitate this multiscale imaging is proposed. The proposed technology will offer greater capability over state-of-the- art products and help further streamline multiscale cellular imaging by simplifying the specimen preparation process and yielding superior imaging performance.