Optical imaging of size, charge, mobility and binding of single proteins

项目总结 蛋白质分析对于理解生命系统中的分子尺度过程、诊断 基于分子生物标记物的疾病研究，以及药物治疗疾病。蛋白质的基本任务 分析包括检测一种蛋白质，鉴定它并确定它与其他蛋白质或分子的相互作用 配基。已经开发了各种技术来执行这些任务，但最不可或缺的是 凝胶和毛细管电泳法、Western Blot(WB)法和酶联免疫吸附试验(ELISA)法。这些 技术根据蛋白质的电荷、大小和与抗体的特定结合来分离和识别蛋白质。 对于分子相互作用分析，目前流行的是表面等离子体共振等检测技术 选择。尽管这些平台在研究实验室和行业中无处不在，但必须结合起来才能提供 完整的蛋白质分析，这是复杂和耗时的。此外，它们缺乏单分子 研究异质过程和实现精确诊断所需的分析能力； 尤其是对于小批量样品。本项目的目标是开发一种能够执行以下任务的检测平台 以上技术的关键功能是具有单分子检测能力。建议的技术 成像没有标记的单个蛋白质，同时测量每个蛋白质的大小、电荷和流动性， 根据其与抗体的特异性结合来识别蛋白质，并量化其与其他蛋白质的相互作用 实时的。亚利桑那州立大学生物电子和生物传感器生物设计中心的团队已经进行了 进行了大量的实验，以验证所提出的技术。在这个R01项目中，团队将解决 剩余的技术挑战，构建一个完整的原型，并验证它在单个 细胞。

PROJECT SUMMARY Protein analysis is essential to the understanding of molecular scale processes in living systems, the diagnosis of diseases based on molecular biomarkers, and the treatment of diseases with drugs. The basic tasks of protein analysis include detecting a protein, identifying it and determining its interactions with other proteins or molecular ligands. Various technologies have been developed to perform these tasks, but the most indispensable ones are gel and capillary electrophoresis, Western Blot (WB) and enzyme linked immunosorbent assay (ELISA). These technologies separate and identify proteins based on a protein’s charge, size, and specific binding to antibodies. For molecular interaction analysis, surface plasmon resonance and other detection technologies are the current choices. Although ubiquitous in both research labs and industry, these platforms must be combined to provide complete analysis of proteins, which is complicated and time consuming. In addition, they lack single molecule analysis capability required for studying heterogenous processes and for achieving precision diagnosis, especially for low volume samples. The present project aims to develop one detection platform that can perform the key functions of the above technologies with single molecule detection capability. The proposed technology images single proteins without labels, measures the size, charge and mobility of each protein simultaneously, identifies the protein based on its specific binding to antibodies, and quantifies its interactions with other proteins in real time. The team at the Biodesign Center for Bioelectronics and Biosensors, ASU, has carried out substantial experiments to demonstrate the proposed technology. In this R01 project, the team will address remaining technical challenges, build a complete prototype and validate it for single protein analysis on single cells.