针对目前循环肿瘤外泌体富集方法中存在的特异性抗体缺乏、抗体易失活、难以重复使用等问题，以提高基于固相材料的外泌体富集检测方法的选择性、灵敏度、回收率、准确度、重现性的需求为切入点，本项目以多种亲和配基的协同亲和作用为特异性识别前提，基于水溶性pH响应聚合物制备多位点识别人工抗体材料。通过构建人工抗体吸附外泌体的动力学和热力学模型，揭示其识别外泌体的机理。通过该材料在肺癌患者体液中的应用，实现外泌体的高效捕获和蛋白质组学研究。本研究结合水溶性pH响应聚合物良好的pH响应特性、水溶性、以及表面功能团易于修饰等特点，发展的均相富集策略可有效避免固相材料存在的相界面和富集不均一性。该材料可代替生物抗体与ELISA等相结合实现外泌体的定性及定量分析，亦可作为富集分离材料应用于生物样品预处理。本研究为发展新的外泌体富集分离方法提供新思路，其在外泌体蛋白质组学中的应用研究为肺癌临床预警和诊治提供重要依据

The efficient capture of circulating tumor exosomes is challenged by the shortage of the specific antibody, the high incidence of antibody to be denatured and lose activities, the difficulty to reuse antibody. In order to improve the selectivity, sensitivity, recovery, accuracy and reproducibility of solid-phase affinity methods, in this project, the synergistic interactions between multiple affinity ligands and exosomes were employed to selectively trap exosomes, and a water-soluble pH-responsive artificial antibody, based on multiple recognition mechanism will be prepared. The mechanism on the recognition of the artificial antibodies towards the exosomes will be studied by the exosomes adsorption thermodynamics and kinetics on the nanomaterials. The application study on biofluids from the lung cancer patients will be conducted to achieve the efficient capture and proteome profiling of exosomes. The pH-responsive polymer exhibits reversible self-assembly and phase separation under pH variations, excellent solubility, controlled surface functionalities. The homogeneous capture strategy supported by the pH-responsive polymer could obtain an outstanding sensitivity, reproducibility and high recovery, compared with biased and unreproducible solid phase based approach. This artificial antibody nanomaterial could be used as the alternatives of biological antibodies, to combine with ELISA for qualitative and quantitative assessment of exosomes, which also could be explored to trap exosomes in biological sample as the sample pretreatment material. This project could provide new insight into developing new isolation methods for exosomes. Its application in proteome profiling of exosomes is of great significance for the clinical early diagnosis and treatment of lung cancer.