TWO-PHOTON SINGLE PARTICLE TRACKING IN THREE DIMENSIONS

了解输运过程的动力学在 生物和医学。在细胞层面上，重要的例子 运输过程包括胞外蛋白的内吞作用， 有丝分裂期间细胞器的运输，抗原物质的吞噬， 核浆转运、病毒对接和感染。 在复杂的三维环境中贩卖人口是一种共享 共同的主题。这些贩运过程很少是被动的或 在蜂窝系统中扩散受控制，但通过活动引导 机制包括分子马达和离子泵。这些 复杂的事件序列很难在以下情况下解决 许多单元格是异步平均的。单粒子跟踪(SPT) S是在20世纪80年代初为解决这一问题而开发的，并已 事实证明，这是一种强大的技术，可以加深我们对 膜蛋白扩散、膜区划和 蛋白质-细胞骨架相互作用。然而，传统的SPT方法 它为二维系统的研究提供了广阔的领域。在这 提案中，申请人计划开发一种3-D粒子跟踪 利用固有的亚飞升定位技术 光子激发。使用实时反馈系统，他可以 动态定位此激励体积以跟随 通过最大化检测到的荧光来传输处于运输中的荧光颗粒 强度。已经很好地确定了细胞的序列 运输通常由生化信号控制。荧光 在显微镜环境中应用光谱方法可以提供 对局部细胞生化状态进行敏感和特定的采样。 苏博士建议将波长分辨光谱学与 3D-SPT连续监测沿线颗粒物微环境 它的蒸腾路径。该仪器的第一个应用将是 重点研究受体介导的蛋白内吞作用。

An understanding of the kinetics of transport processes is crucial in biology and medicine. On the cellular level, examples of important transport processes include endocytosis of extracellular protein, organelle transport during mitosis, phagocytosis of antigen material, mRNA nucleo-cytoplasmic transport, virus docking and infection. Trafficking inside a complex three-dimension environment is a shared common theme. These trafficking processes are rarely passive or diffusion-controlled in cellular system but are guided through active mechanisms including molecular motors and ion pumps. These complex sequences of events are difficult to resolve when the action of many cells are asynchronously averages. Singe particle tracking (SPT) was developed in the early 1980's to address this problem and has proved to be a powerful technique to further our understanding of membrane protein diffusion, membrane compartmentalization and protein-cytoskeleton interaction. However, the traditional SPT method which uses wide field to the study of two-dimensional systems. In this proposal, the applicant plans to develop a 3-D particle tracking technique utilizing the inherent sub-femtoliter localization of two photon excitation. Using a real-time feedback system, he can dynamically position this excitation volume to follow the position of a fluorescent particle under transport by maximizing detected fluorescent intensity. It has been well established that the sequence of cellular transport are often controlled by biochemical signals. Fluorescent spectroscopic method applied in microscopy setting can provide sensitive and specific sampling of the local cellular biochemical states. Dr. So proposes to integrate wavelength resolved spectroscopy with 3D-SPT to continuously monitor the particle micro- environment along its transpiration path. The first application of this instrument will be focused on the study of receptor-mediate protein endocytosis.