VIRTUAL FCS

这个子项目是利用由NIH/NCRR资助的中心拨款提供的资源的许多研究子项目之一。子项目和调查员(PI)可能从另一个NIH来源获得了主要资金，因此可能会出现在其他CRISE条目中。列出的机构是针对中心的，而不一定是针对调查员的机构。荧光相关光谱(FCS)基于对荧光粒子在飞升观测体积中移动时发出的光子的统计分析，提供了关于分子迁移率、浓度、分子络合物组成、解离常数和反应动力学的定量信息。已建立的FCS理论基于粒子的自由扩散以及参与某些类型的反应的粒子来预测光子到达时间的统计。然而，对于活细胞中发现的更复杂的颗粒系统，包括膜结构、细胞骨架、与多个伙伴的相互作用和分子拥挤，没有明确的解决方案。为了解释从活细胞获得的FCS数据，将使用空间蒙特卡罗方法对体内环境进行建模。目前，我们正在开发模拟单个粒子在反射反应体积内随机扩散的软件。粒子可以具有不同的扩散系数和荧光量子产率，并可以进行不同的一级和二级反应。可以在反应体积内限定各种反射面。观测量被定义为空间亮度函数。光子发射概率取决于粒子相对于观察体中心的位置。仿真结果被用于验证在体FCS数据的数据分析方法。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Fluorescence correlation spectroscopy (FCS) based on statistical analysis of photons emitted by fluorescent particles as they move through a femtoliter observation volume provides quantitative information about mobility of molecules, concentration, composition of molecular complexes, dissociation constants, and reaction kinetics. The established theory of FCS predicts the statistics of photon arrival times based on free diffusion of particles, and particles involved in certain kinds of reactions. However explicit solutions do not exist for more complex particle systems found in live cells and involving membrane structures, cytoskeleton, interactions with multiple partners, and molecular crowdedness. To interpret the FCS data obtained from live cells, the in vivo environment will be modeled by spatial Monte Carlo method. Currently we are developing software that models individual particles randomly diffusing inside a reflective reaction volume. Particles can have different diffusion coefficient and fluorescence quantum yield and can engage in different first and second order reactions. Various reflective surfaces can be defined inside the reaction volume. The observation volumes is defined as a spatial brightness function. Photon emission probability depends on particle position relative to the center of the observation volume. Simulation results are used to validate data analysis methods for in vivo FCS data.