Development and Commercialization of a Sample Preparation System for Time Resolved Cryo-Electron Microscopy

时间分辨冷冻电子显微镜样品制备系统的开发和商业化

• 批准号: 10231377
• 负责人: JOACHIM FRANK
• 金额: $ 84.04万
• 依托单位: HUMMINGBIRD PRECISION MACHINE COMPANY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2023-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10231377
• 关键词: Biological; Biological Process; Cone; Consumption; Cost efficiency; Cryoelectron Microscopy; Data; Development; Dimensions; Documentation; Electron Beam; Electrons; Environmental Monitoring; Ethane; Evolution; Freezing; Gases; Humidity; Hydration status; Image; Imaging Techniques; Kinetics; Liquid substance; Measures; Methods; Microfluidics; Molecular Structure; Pathway interactions; Phase; Population; Population Distributions; Positioning Attribute; Preparation; Process; Production; Reaction; Reaction Time; Recipe; Reproducibility; Research; Research Personnel; Resolution; Running; Sampling; Small Business Technology Transfer Research; Speed; Standardization; Structure; System; Techniques; Temperature; Testing; Time; Universities; Work; X-Ray Crystallography; biological research; biological systems; commercialization; cost; design; experimental study; improved; millisecond; operation; particle; preservation; pressure; prototype; software development; temporal measurement; touchscreen; user-friendly

Cryo-electron microscopy (cryo-EM) is a powerful technique for imaging biological molecules in which samples are preserved in a frozen, hydrated state before imaging using an electron beam. The technique provides an alternative to X-ray crystallography for structural characterization of biological samples that are not crystallizable or do not crystallize in their native state. While cryo- EM sample preparation technique has become more efficient in recent years with the development of automatic sample vitrification systems to perform grid preparation, the field of cryo-EM is hindered by the inability to perform time-resolved experiments on a sufficiently fast time scale to observe the progression of many biological reactions due to the inherent lag of traditional blotting methods. The Frank group at Columbia University has developed a lab- prototype of a system to enable this missing capability and enables millisecond time control over the reaction time of sampling biological reactions in the grid preparation process. The purpose of this Fast-Track STTR proposal is to develop the time-resolved system built by the Frank group into a commercial production unit and launch it as a product. Initial Phase I milestones in the proposed work will focus on building a prototype with added control of reaction parameters to determine necessary control variables in the system and to optimize sample grid preparation and time resolution. Further work in Phase II of this Fast-Track proposal will create a streamlined, user-friendly production unit which will maintain the necessary controls as determined in initial milestones, while optimizing manufacturing time and reducing overall production costs. The resulting system will add another dimension to the field of cryo-EM, providing users with the capability to capture transient intermediates in reaction progression studies and further clarifying proposed reaction mechanisms with time resolution.

冷冻电子显微镜（cryo-EM）是一种强大的生物分子成像技术， 该样品在使用电子束成像之前以冷冻、水合状态保存。 该技术提供了一种替代X射线晶体学的结构表征， 不可结晶或在其天然状态下不结晶的生物样品。当冷冻- EM样品制备技术近年来变得更加有效， 自动样品玻璃化系统的开发，以执行网格制备， cryo-EM由于不能在足够快的时间上进行时间分辨实验而受到阻碍。 时间尺度来观察许多生物反应的进展，由于固有的滞后， 传统的印迹方法。哥伦比亚大学的弗兰克小组开发了一个实验室- 一个系统的原型，使这一缺失的能力，并使毫秒时间控制， 网格制备过程中采样生物反应的反应时间。 这一快速通道STTR提案的目的是开发由 弗兰克把它分成一个商业生产单位，并把它作为一种产品推出。初始阶段I 拟议工作的里程碑将集中在建立一个原型，增加了反应控制 参数，以确定系统中必要的控制变量，并优化样品网格 准备和时间分辨率。该快速通道提案第二阶段的进一步工作将建立一个 精简、方便用户的生产单位，将保持必要的控制， 在最初的里程碑中确定，同时优化制造时间， 生产成本由此产生的系统将为冷冻EM领域增加另一个维度， 为用户提供捕获反应过程中瞬时中间体的能力 研究和进一步澄清提出的反应机制与时间分辨率。