IMAGING BARRIERS TO MOLECULAR DIFFUSION BY PAIR CORRELATION FUNCTIONS

通过成对相关函数对分子扩散的障碍进行成像

• 批准号: 8365762
• 负责人: ENRICO GRATTON
• 金额: $ 4.27万
• 依托单位: UNIVERSITY OF CALIFORNIA-IRVINE
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-08-20 至 2012-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8365762
• 关键词: Cells; Complex; Diffusion; Event; Fluorescence; Funding; Grant; Image; Kinetics; Label; Laboratories; Lasers; Life; Location; Measurement; Measures; Modeling; Molecular; National Center for Research Resources; Nuclear Envelope; Nuclear Pore; Principal Investigator; Procedures; Research; Research Infrastructure; Resources; Source; Techniques; Time; Transport Process; United States National Institutes of Health; base; cost; novel; single molecule

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. I study the intracellular diffusivity of a model substrate (NLS-GFP) by using a novel and unconventional fluorescence correlation technique, based on pair correlation functions. I detect molecular transport events by measuring the time a single molecule takes to move from a specific location to another within the cell. Here is the novelty with respect to the traditional FCS approach that measures the time a molecule takes to cross the laser beam. By this approach obstacle to diffusion can be detected both within cellular compartments (restrictions to free diffusion) and across the nuclear envelope barrier (restrictions due to the presence of nuclear pore channels). In both cases the measurement affords a quantitative description of the kinetic steps of NLS-GFP transport at the single molecule level, and in living cells, without the need for complex labeling procedures and cell manipulations.

该子项目是利用资源的众多研究子项目之一 由 NIH/NCRR 资助的中心拨款提供。子项目的主要支持 并且子项目的首席研究员可能是由其他来源提供的， 包括其他 NIH 来源。 子项目可能列出的总成本 代表子项目使用的中心基础设施的估计数量， NCRR 赠款不直接向子项目或子项目工作人员提供资金。 我通过使用基于配对相关函数的新颖且非常规的荧光相关技术来研究模型底物 (NLS-GFP) 的细胞内扩散率。我通过测量单个分子从细胞内的特定位置移动到另一个位置所需的时间来检测分子运输事件。这是相对于传统 FCS 方法的新颖之处，传统的 FCS 方法测量分子穿过激光束所需的时间。通过这种方法，可以在细胞区室（自由扩散的限制）和跨核膜屏障（由于核孔通道的存在而限制）内检测扩散障碍。在这两种情况下，测量都可以定量描述单分子水平和活细胞中 NLS-GFP 转运的动力学步骤，而不需要复杂的标记程序和细胞操作。