内标比率型SERS探针的构建及外泌体多组分分析检测研究

Exosomes are small nano-sized vesicles secreted by most living cells, the type and expression level of surface protein biomarkers are strongly correlated with the presence and progression of a certain cancer class. Exosomes are one of the most promising cancer markers in the field of “liquid biopsy”. However, the traditional analysis methods are labor-intensive, time-consuming and have poor specificity. This project aims to design and prepare surface enhanced Raman scattering probes with abundant “hot spots”, and construct a ratiometric analysis platform based on the aptamer directed “core-satellite” structures. Herein, unique nanostructures and suitable sensing interfaces are designed by taking advantages of the controllability and surface plasmon resonance characteristics of nanomaterials; in addition, specific aptamers are introduced to achieve single-cell source exosome multi-component analysis and multi-cell source exosome simultaneous analysis. The mechanism of exosome-mediated probe structure changes will be investigated, and the conversion and amplification mechanism of surface enhanced Raman scattering signals will be emphatically clarified. The successful implementation of this project will promote the further development of surface enhanced Raman probes in the field of analysis, provide new theoretical and technical support for the analysis of exosomes. It is of great significance to promote the clinical utility of exosomes for cancer diagnostics.

外泌体是由多种活细胞分泌的纳米级囊泡小体，其表面蛋白标志物的表达水平和类型与癌症的进展和类别密切相关，具有非常重要的研究价值。然而，传统的外泌体分离检测方法，前处理复杂、耗时、特异性差。本项目拟设计制备具有丰富“热点”的表面增强拉曼探针，通过碱基互补配对获得“核-卫星”组装体结构，构建基于表面增强拉曼散射的内标比率型快速分析检测平台。借助纳米材料的可调控性和表面等离子共振特性，设计独特的纳米结构和适宜的传感界面；结合特异性识别核酸适配体，实现单细胞源外泌体多组分分析和多细胞源外泌体同时分析检测。研究靶标外泌体介导探针组装结构变化机理，阐明其对表面增强拉曼散射信号的转换与放大机制。本项目的顺利实施将促进表面增强拉曼探针在分析检测领域的进一步发展，为外泌体的分析检测提供新的理论和技术支持，对推进外泌体在癌症诊断领域中的实用化进程具有重要意义。

外泌体是由多种活细胞分泌的纳米级囊泡小体。研究表明，外泌体是“液体活检”领域最有前途的癌症标志物之一。外泌体表面蛋白标志物具有显著的细胞来源特性，它们的类型和表达水平与某些癌症类别的存在和进展密切相关。如果可以实现外泌体水平和标志分子的灵敏、快速分析检测，就有可能实时监测体内循环体液甚至组织微环境的分子水平改变，做到提前“预知”癌症。然而，生物体液基质复杂，除了外泌体，还有许多复杂的背景干扰，如细胞碎片、蛋白以及其他种类的囊泡等。传统的外泌体分离检测方法，前处理复杂、耗时、特异性差、检测灵敏度低，要实现外泌体的准确分析检测仍然面临着极大地挑战。.基于以上背景，制备了内标比率型表面增强拉曼探针，进行信号的自我校正，以及通过识别外泌体多类型识别位点等策略降低体液基质复杂的背景干扰；制备了具有丰富“热点”的金纳米花探针，设计了无酶信号放大策略以提高检测的灵敏度；并利用磁分离技术实现外泌体的分离和富集。该项目所构建的分离检测方法，对多种类细胞来源外泌体具有通用性。本项目的完成，将为外泌体准确检测和疾病早期诊断提供有价值的理论依据和技术支持。