Development of Room-Temperature Storage Technique for Plasma/Serum Biospecimens

血浆/血清生物样本室温储存技术的发展

• 批准号: 8431941
• 负责人: Alptekin Aksan
• 金额: $ 22.71万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-09-17 至 2015-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8431941
• 关键词: Adsorption; Ascites; Biochemical Reaction; Biological Markers; Biological Preservation; Bronchoalveolar Lavage Fluid; Characteristics; Chemicals; Cold Chains; Collection; DNA; Desiccation; Development; Diagnostic; Epidemiology; Equipment; Farming environment; Freezing; Future; Genomics; Glass; Goals; Hour; Hydration status; Ice; Kinetics; Lipids; Liquid substance; Malignant Neoplasms; Medicine; Methodology; Methods; Outcome; Paper; Plasma; Porifera; Process; Proteins; Proteomics; Protocols documentation; RNA; Research; Research Project Grants; Saliva; Sampling; Serum; Serum Proteins; Site; Specimen; Stress; Structure; Techniques; Technology; Temperature; Testing; Therapeutic; Time; United States National Institutes of Health; Urine; Weight; biobank; cost; cryogenics; experience; improved; innovation; macromolecule; metabolomics; novel; research study; success; sugar; tool

DESCRIPTION (provided by applicant): Currently, millions of serum biospecimens are being stored in biorepositories across the nation, while tens of thousands of new biospecimens are added to the pool daily. These biospecimens are stored for future research, mainly for proteinaceous biomarker discovery and verification (e.g. for diagnostic, therapeutic, and epidemiologic outcomes). The success of biomarker research not only depends upon the availability of the tools (proteomic, peptidomic, lipidomic and metabolomic technologies) to extract information from biospecimens, but also on the availability of "high quality" biospecimens. However, in most biorepositories, serum is stored by freezing without following any preservation protocol; the samples are directly placed in -20, -40 or -80oC freezers, in the absence of any cryoprotectant, where they experience very slow cooling (1-2o C/min). It is well known that these conditions impose very harsh chemical and physical stresses on macromolecules, altering their characteristics (structure and activity), often irreversibly. Recent evidence has shown that some of the most promising proteinaceous cancer biomarkers are indeed very susceptible to freeze/thaw and frozen state storage. Therefore, it is plausible that frozen state storage may cause most of the potential biomarker information in the stored sera biospecimens to be lost forever. Our long-term goal is to eliminate the requirement for frozen state storage and develop the techniques to store serum biospecimens at room temperature using isothermal vitrification technology. Isothermal vitrification is the process by which liquids doped with sugars are desiccated to a "glass" (a very viscous fluid). In this state, biochemical reactions are halted, degradation of the specimen is stopped, and macromolecules are stabilized in their native states. Isothermal vitrification will eliminate the exposure of the proteinaceous biomarkers to freeze/thaw stresses and to frozen state storage damage and thus will substantially increase the quality of the stored biospecimens. It will also present a more economical alternative to freezing, since storage of specimens in freezer-farms will no longer be needed. We will accomplish our goal by achieving the following four Aims: Aim #1. Develop a panel of lyoprotectant chemicals to be added to serum samples for isothermal vitrification: Demonstrate that the lyoprotectant cocktail enables rapid and uniform vitrification while stabilizing sera proteins. Aim #2. Determine the retention and elution efficiency of sera proteins following vitrification by adsorption in standard filter paper using the developed lyoprotectant cocktail. Aim #3. Determine whether specific sera proteins are altered following optimized vitrification and elution conditions from the optimal matrix; document protein depletion, aggregation and degradation. Aim #4. Develop and validate an electrospun sponge for streamlined vitrification of serum samples at sera collection sites. PUBLIC HEALTH RELEVANCE: The advent of genomics and proteomics for personalized medicine has placed biospecimen research on the forefront of NIH priorities, since research projects are only as good as the biospecimens used. We are developing methodology to stably store liquid biospecimens, such as sera, at room temperature. This technology will eliminate the need for long-term storage of serum samples in freezer farms, which are very costly to operate.

描述（由申请人提供）：目前，数百万份血清生物标本储存在全国各地的生物储存库中，而每天有数万份新的生物标本添加到池中。这些生物样本被储存用于未来的研究，主要用于蛋白质生物标志物的发现和验证（例如用于诊断、治疗和流行病学结果）。生物标志物研究的成功不仅取决于从生物标本中提取信息的工具（蛋白质组学、肽组学、脂质组学和代谢组学技术）的可用性，而且取决于“高质量”生物标本的可用性。然而，在大多数生物储存库中，血清通过冷冻储存而不遵循任何保存方案;样品直接置于-20、-40或-80 ° C的冷冻机中，在没有任何冷冻保护剂的情况下，它们经历非常缓慢的冷却（1- 2 ° C/min）。众所周知，这些条件对大分子施加了非常苛刻的化学和物理应力，改变了它们的特性（结构和活性），通常是不可逆的。最近 有证据表明，一些最有希望的蛋白质类癌症生物标志物确实对冷冻/解冻和冷冻状态储存非常敏感。因此，冷冻状态储存可能会导致储存的血清生物标本中的大多数潜在生物标志物信息永久丢失，这是合理的。 我们的长期目标是消除对冷冻状态储存的要求，并开发使用等温玻璃化技术在室温下储存血清生物标本的技术。等温玻璃化是液体 掺杂有糖的液体被干燥成“玻璃”（一种非常粘稠的液体）。在这种状态下，生化反应停止，标本的降解停止，大分子稳定在其天然状态。等温玻璃化将消除蛋白质生物标志物暴露于冷冻/解冻应力和冷冻状态储存损伤，因此将显著提高储存的生物标本的质量。它也将提供一种比冷冻更经济的替代方法，因为不再需要在冷冻场储存标本。我们将通过实现以下四个目标来实现我们的目标：目标#1。开发一组冻干保护剂化学品，添加到血清样品中进行等温玻璃化：证明冻干保护剂混合物能够快速均匀地玻璃化，同时稳定血清蛋白。 目标2。测定血清蛋白的保留和洗脱效率 随后通过使用开发的冻干保护剂混合物在标准滤纸中吸附进行玻璃化。 目标3。确定特定血清蛋白在优化玻璃化和最佳基质洗脱条件后是否发生改变;记录蛋白质消耗、聚集和降解。 目标4。开发并验证静电纺丝海绵，用于在血清采集点对血清样品进行流线型玻璃化冷冻。 公共卫生关系：个性化医疗的基因组学和蛋白质组学的出现使生物标本研究成为NIH优先考虑的问题，因为研究项目的好坏取决于所使用的生物标本。我们正在开发在室温下稳定储存液体生物标本（如血清）的方法。这项技术将消除在冷冻农场长期储存血清样本的需要，因为冷冻农场的运营成本非常高。