Method and Device for Rodent Germplasm Vitrification

啮齿类种质玻璃化冷冻方法及装置

• 批准号: 7800253
• 负责人: Hongbin Ma
• 金额: $ 18.5万
• 依托单位: UNIVERSITY OF MISSOURI-COLUMBIA
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-04-15 至 2012-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7800253
• 关键词: Acrosome; Area; Binding; Biomedical Research; Caliber; Cell Survival; Cells; Characteristics; Chemicals; Complex; Cryopreservation; Cryoprotective Agents; Crystal Formation; Device Designs; Devices; Dimensions; Dimethyl Sulfoxide; Embryo; Engineering; Excision; Family suidae; Film; Glycerol; Goals; Heating; Ice; Investigation; Liquid substance; Measures; Medicine; Membrane; Metals; Methods; Microfabrication; Microfilaments; Modeling; Modification; Morphology; Mus; Nitrogen; Performance; Porosity; Procedures; Process; Pump; Reproductive Medicine; Research; Research Infrastructure; Research Personnel; Resources; Rodent; Silicon; Sperm Motility; Structure; Surface; Survival Rate; Suspension substance; Suspensions; System; Techniques; Temperature; Testing; Thick; Time; Toxic effect; Work; base; cell injury; cell type; cost; cryogenics; design; efficacy testing; evaporation; gene therapy; improved; melting; novel; oncology; operation; physical model; prevent; public health relevance; research study; sperm cell

DESCRIPTION (provided by applicant): The over-all goal of this application is to establish a universal and easy-to-use germplasm cryopreservation approach by developing a novel ultra-fast cooling device system. This device system achieves vitrification of germplasm suspensions without any cryoprotective agent (CPA) or with low concentration CPA (<1M). We will do this by combining cutting-edge engineering techniques including: (1) cryogenic thin film evaporation (TFE), (2) thermal fins, and (3) microfabrication techniques. Germplasm cryopreservation is essential to many areas of medicine such as reproductive medicine, oncology, and gene therapy. However, current cryopreservation methods and devices are very different for different types of germplasms or even within cell types and they are sometimes very difficult to use. Due to the complications and inefficiencies involved in these methods, many cells types cannot be successfully cryopreserved at all and many others respond very poorly to current methods (i.e., more than 50% of the cells are lost during the process). Therefore, it is important to establish a universal, efficient cell cryopreservation method and develop corresponding devices that require relatively simple operations. Direct vitrification (solidification without crystal formation) of cell suspensions without permeating CPAs (e.g. glycerol and DMSO), or with relatively low concentrations (<1M) of CPAs, is suitable for cryopreservation of essentially all cell types. This is the case because (1) vitrification prevents cell damages caused by ice formation; (2) a low concentration of CPA significantly lowers CPA toxicity and osmotic damages during CPA addition and removal procedures. However, vitrification with low concentration CPAs requires ultra-fast cooling rates (106-7K/min). Since none of the current cryopreservation methods and devices can achieve these ultra-fast cooling/warming rates, successful completion of this work will significantly enhance the underlying infrastructure for many areas of biomedical research and save a significant amount of resources by providing a universal, effective cell cryopreservation system. This work will be done by completing the following specific aims: (1) theoretical and experimental investigations on applications of the three cutting-edge techniques in cryopreservation, (2) determination of the optimal design of the proposed device system based on the above investigations, and (3) empirical testing the device system by cell survival rates. PUBLIC HEALTH RELEVANCE (provided by applicant): Germplasm cryopreservation is essential to many areas of medicine such as reproductive medicine, oncology, and gene therapy. However, current cryopreservation methods and devices are very different for different types of germplasms or even within cell types and they are sometimes very difficult to use. The over-all goal of this application is to establish a universal and easy-to-use germplasm cryopreservation approach by developing a novel ultra-fast cooling device system.

描述（由申请人提供）：此应用程序的所有目标是通过开发一种新型的超快速冷却设备系统来建立通用且易于使用的种质冷冻保存方法。该设备系统可实现玻璃质的玻璃质悬浮液，而无需任何 冷冻保护剂（CPA）或低浓度CPA（<1m）。我们将通过结合最先进的工程技术来做到这一点，包括：（1）低温薄膜蒸发（TFE），（2）热鳍和（3）微加工技术。种质冷冻保存对于许多医学领域，例如生殖医学，肿瘤学和基因疗法至关重要。但是，对于不同类型的种系甚至在细胞类型中，当前的冷冻保存方法和设备截然不同，有时很难使用它们。由于这些方法中涉及的并发症和效率低下，因此许多细胞类型无法成功保存，因此许多其他细胞对当前方法的反应很差（即，在此过程中丢失了50％以上的细胞）。因此，建立一种通用，有效的细胞冷冻保存方法并开发需要相对简单操作的相应设备很重要。细胞悬浮液的直接玻璃化（没有晶体形成），而无需渗透CPA（例如甘油和DMSO）或CPA浓度相对较低的浓度（<1m），适用于基本所有细胞类型的冷冻保存。就是这样，因为（1）玻璃化阻止了由冰的形成引起的细胞损伤； （2）低浓度的CPA显着降低了CPA的毒性和渗透损害，在CPA添加和去除程序期间。但是，低浓度CPA的玻璃化需要超快速冷却速率（106-7k/min）。由于当前的冷冻保存方法和设备都无法实现这些超快速的冷却/变暖率，因此成功完成这项工作将显着增强许多生物医学研究领域的基础基础设施，并通过提供通用的有效的，有效的细胞冷冻保存系统来节省大量资源。这项工作将通过完成以下特定目的完成：（1）对三种尖端技术在冷冻保存中的应用的理论和实验研究，（2）基于上述研究确定所提出的设备系统的最佳设计，（3）通过细胞存活率来实现设备系统的经验测试。 公共卫生相关性（由申请人提供）：种质冷冻保存对于许多医学领域，例如生殖医学，肿瘤学和基因疗法至关重要。但是，对于不同类型的种系甚至在细胞类型中，当前的冷冻保存方法和设备截然不同，有时很难使用它们。该应用程序的所有目标是通过开发一种新型的超快速冷却设备系统来建立通用且易于使用的种质冷冻保存方法。