Detachable Nozzle Piezoelectric Dispense System for Cryo-TEM Sample Preparation

描述(由申请人提供)：项目摘要/摘要工程技术(EA)建议开发一种可拆卸的喷嘴压电分配系统，用于低温透射电子显微镜(Cryo-TEM)样品制备仪器。低温电子显微镜是研究天然水相生物大分子结构的重要工具。为低温透射电子显微镜制备“薄冰”冷冻水样的过程具有挑战性且费时费力，几十年来基本上没有变化。通常，将单个含水生物样品(~3 L)手动涂抹在薄的多孔电子显微镜(EM)网格上，然后将网格吸去多余的样品，然后将其插入液体乙烷中进行闪速冷冻。这一过程有望在网格上留下足够的区域，使其具有合适的玻璃化“薄冰”，通常厚度为50到200纳米，这是电磁成像所需的。本文提出的可拆卸式压电喷嘴点胶系统是一项关键的创新，将导致低温瞬变电子显微镜工作流程的生产率、质量和能力的突破。低温瞬变电子显微镜工艺的主要优点如下：1.在每个低温瞬变电子显微镜网格上应用多种不同的样品(或样品条件)。2.提高工艺控制和可接受的薄冰产量，超过当前的标准印迹方法。3.较低的起始样本量要求和单位网格消耗。4.允许对多个样品的混合/结合事件进行时间分辨研究。EA将设计、制造和优化一种多通道可拆卸喷嘴压电分配系统，该系统能够将几种不同生物样本(或样本条件)的液滴(小于50皮升)分配到每个透射电子显微镜网格上。NRAMM(国家自动化分子显微镜资源，斯克里普斯，La Jolla CA)将三通道可拆卸喷嘴压电点样系统改装成定制的Cryo-TEM样品制备仪(Spotiton仪器平台)。用于测试、优化和验证。

DESCRIPTION (provided by applicant): Project Summary / Abstract Engineering Arts (EA) proposes the development of a detachable nozzle piezoelectric dispense system for use in Cryo-TEM (Cold - Transmission Electron Microscopy) sample preparation instrumentation. Cryo-TEM is an important tool for studying the structure of biological macromolecules in their native aqueous state. The challenging and laborious process of preparing 'thin-ice' frozen aqueous samples for Cryo-TEM has remained essentially unchanged for decades. Typically a single aqueous biological sample (~3¿l) is manually applied onto a thin porous Electron Microscopy (EM) grid which is then blotted to remove excess sample and then plunged into liquid-ethane for flash freezing. This process hopefully leaves enough regions on the grid with the proper vitrified 'thin-ice', typically 50 to 200nm thickness, required for EM imaging. The detachable piezoelectric nozzle dispense system proposed here is a key innovation that will lead to productivity, quality and capability breakthroughs for Cryo-TEM workflow. The following key benifits for Cryo-TEM process will be realized: 1. Apply many different samples (or sample conditions) to each Cryo-TEM grid. 2. Increase process control and yields of acceptable thin-ice over current standard blotting methods. 3. Lower starting sample volume requirements and consumption per grid. 4. Allow time-resolved study of mixing / binding events of multiple samples. EA will design, fabricate and optimize a multichannel detachable nozzle piezoelectric dispense system that is capable of dispensing droplets (less than 50 picoliters) of several different biological samples (or sample conditions) onto each TEM grid. A three channel detachable nozzle piezoelectric dispense system retrofitted onto a custom Cryo-TEM sample preparation instrument (Spotiton instrument platform) at NRAMM (National Resource for Automated Molecular Microscopy, Scripps, La Jolla CA.) for testing, optimization and validation.