High Throughput Extracellular Flux Analyzer

描述(由申请人提供)： 在动物细胞中有两条途径产生大量的ATP--线粒体氧化磷酸化和糖酵解。生物学家对这些ATP产生过程的调控非常感兴趣，但这些过程很难在细胞系、原代细胞或组织样本中进行研究，特别是在适合高通量的96孔结构中。最近，海马生物科学公司开发了XF24胞外通量分析仪，可以连续测量在24孔板中培养的细胞的氧气消耗和酸形成。通量分析仪使研究人员能够重复和非破坏性地测量单细胞培养中的氧化磷酸化、糖酵解、营养利用和其他过程。然而，24孔仪器不能为大型化学库的初步筛选提供足够的吞吐量。因为原代细胞或组织需要在有限的时间内进行检查，所以使用这些样本的研究人员大大限制了可以对给定动物进行检查的变量的数量。因此，我们的具体目标是1)获得一台用于测量氧气摄取和酸释放的海马氏96孔胞外通量分析仪，以及2)将该仪器置于多用户环境中，为个别用户项目提供技术支持、用户培训和应用专业知识。这项新兴技术将独特地允许NIH资助的研究人员从事各种项目，以评估正常生理和疾病模型中的生物能量学和代谢反应，包括癌症、酒精性肝毒性、黄斑变性、心脏缺血和衰竭、多器官衰竭和糖尿病。这些研究的结果可能会导致为其中一些疾病开发新的治疗方法。

DESCRIPTION (provided by applicant): Two pathways generate the bulk of ATP in animal cells -mitochondrial oxidative phosphorylation and glycolysis. Regulation of these ATP-generating processes is of fundamental interest to biologists, but these processes are difficult to study in cell lines, primary cells or tissue samples, especially in a 96-well configuration amenable to high throughput. Recently, Seahorse Bioscience has developed the XF24 Extracellular Flux Analyzer that permits serial measurements of O2 consumption and acid formation of cells cultured in 24-well plates. The flux analyzer enabled investigators to measure repetitively and non-destructively oxidative phosphorylation, glycolysis, nutrient utilization and other processes in single cell cultures. However, the 24-well instrument does not offer sufficient throughput for primary screening of large chemical libraries. Because primary cells or tissues need to examined in a limited time period, investigators using these samples are significantly limited to the number of variables that can be examined for a given animal. Accordingly, our specific aims are to 1) acquire a Seahorse 96-well Extracellular Flux Analyzer for measurement of O2 uptake and acid release, and 2) place this instrument in a multi-user environment that provides technical support, user training and applications expertise in support of individual user projects. This emerging technology will uniquely permit NIH-funded researchers working on a variety of projects to assess bioenergetics and metabolic responses in normal physiology and models of disease, including cancer, alcoholic hepatotoxicity, macular degeneration, heart ischemia and failure, multiple organ failure and diabetes. The results of those studies could lead to the development of new therapeutic treatments for some of these diseases.