REAL-TIME SINGLE-MOLECULE CHEMICAL MICROSCOPY

描述(改编自申请人的摘要)：本研究项目 旨在开发和应用尖端分析化学技术和 单分子生物传感器(SMB)和单分子生物传感器 用于实时跟踪一系列生物事件的化学显微镜 (例如，T细胞免疫反应)在SM级别实时进行。的研究。 热力学(亲和力、化学计量)和动力学(机理、速率常数) 单细胞表面配体-受体结合反应参数的研究 在SM层面上是了解发病的关键一步 生物级联反应、疾病感染和潜在药物的设计。 这些创新的方法将证明其在解开 目前阻碍了我们对癌症等疾病的完全了解 和艾滋病从发病和发展到诊断和治疗 治疗。这些研究也将提供独特的机会来理解 SM级别的基本理论(例如，锁和钥匙模型)。最终， 这项研究可能导致发现新的细胞信号通路和 生物医学研究新技术的发明。本发明 以及生物医学用新型分子器件(如SMB)的发展 应用程序也将提出新的挑战，并开辟新的 生物医学工程师的机会。对总体方案进行了答复 最先进的超灵敏分析和单次扫描的新方法。 分子检测(SMD)最近在我们的 实验室。两个相互关联的研究目标是：1)开发新型SMB和 SM化学显微镜用于实时跟踪单细胞免疫反应。 将使用SMB与SM的实时耦合来构建工具 荧光显微镜。这台仪器将能够确定 细胞信号的实时热力学和动力学分布 细胞表面与单个配体-受体相互作用的途径 单细胞，以及2)在单细胞和SM水平上实时演示 监测T细胞免疫反应的调节。中小型企业将用于 在单个T细胞表面触发单个配体-受体结合事件 手机。相关的细胞信号通路(例如，水解酶 肌醇磷脂在质膜中，在细胞内增加 第二信使水平)将使用SM化学公司进行实时监测 显微镜。

DESCRIPTION (Adapted from the applicant's abstract): This research project aims to develop and apply cutting-edge analytical chemistry technologies and methodologies such as single-molecule biosensors (SMB) and single-molecule chemical microscopy for real-time tracking of a cascade of biological events (e.g., T cell immune response) in real-time at the SM level. The study of thermodynamic (affinity, stoichiometry) and kinetic (mechanism, rate constant) parameters of ligand-receptor binding reactions on the surface of single cells at the SM level is an essential step in gaining an understanding of the onset of biological cascades, disease infection and the design of potential drugs. These innovative approaches will prove value in unraveling mysteries that presently prohibit us from completely understanding diseases such as cancer and AIDS from their onset and development through their diagnosis and treatment. These studies will also provide unique opportunities to understand fundamental theories (e.g., lock-and-key model) at the SM level. Ultimately, this research may lead to the discovery of new cellular signaling pathways and the invention of novel technologies for biomedical research. The invention and development of novel molecule devices (e.g., SMB) for biomedical applications will also raise the new challenges and open up the new opportunities for biomedical engineers. The general scheme is replied upon the novel approaches of state-of-the-art ultrasensitive analysis and single- molecule detection (SMD) recently described and demonstrated in our laboratory. Two interrelated research aims are: 1) To develop novel SMB and SM chemical microscopy for real-time tracking of single-cell immune response. Instrumentation will be constructed using real-time coupling of SMB with SM fluorescence microscopy. This instrumentation will be capable of determining the real-time thermodynamic and kinetic profiles of cellular signaling pathways associated with single ligand-receptor interaction on the surface of single cells, and 2) To demonstrate at the single-cell and SM level, real-time monitoring of the regulation of T cell immune response. A SMB will be used to trigger a single ligand-receptor binding event on the surface of a single T cell. The associated cellular signaling pathways (e.g., hydrolysis of inositol phospholipids in the plasma membrane, increase in intracellular second messenger levels) will be monitored in real-time using SM chemical microscopy.