新型SERS纳米材料构筑及其在细胞内活性氧物种检测中的应用研究

Reactive oxygen species (ROS) play a significant role in intracellular biochemical reaction in the process of aerobic metabolism of living organisms. However, when overproduced, these ROS will cause cell death，thus leading to diseases. Therefore, it is of great importance to develop ROS analysis method for the diagnosis of diseases at a cellular level. This program has been proposed to exploit a novel method for the sensitive and selective determination of intracellular ROS, which is based on the prepared nanoparticles and surface-enhanced Raman scattering (SERS) with advantages of high sensitivity, weak water background, high-speed acquisition and real-time analysis. Molecular assembly and in situ growth were employed to prepare nanoparticles with excellent SERS activity. Then, a series of selected Raman tags which are sensitive to different ROS were modified on the surface of nanoparticles uniformly by interface control and surface modification to construct highly sensitive and selective SERS nanosensors for ROS. The prepared nanosensors will be delivered to the living cells, and permit the concentration of different ROS to be monitored in real-time and on-line, thus establishing a novel method for detection of ROS. The research results are expected to provide new research ideas for understanding of oxidative stress-related diseases at a cellular level, laid the foundation for application of SERS nanosensors.

活性氧物种（ROS）在生物体新陈代谢过程中对细胞内生化反应有操控性作用，但其过量产生会导致细胞凋亡，引发疾病。因此，发展在细胞水平上检测ROS的分析方法对疾病的早期诊断具有重要的意义。本项目拟通过构筑新型纳米材料，结合表面增强拉曼散射（SERS）灵敏度高、水干扰小、检测速度快、可原位分析等优点，开发活细胞内ROS的高选择性、高灵敏检测新方法。项目通过分子组装、原位生长等方法合成增强效果好、稳定性高的纳米材料，筛选特异性响应不同ROS的拉曼标记分子，结合物理或化学修饰等手段将拉曼标记分子均匀地组装在纳米材料表面，构建高灵敏高特异性识别ROS的新型SERS纳米传感器；在此基础上，对细胞内ROS进行实时、在线原位的定性、定量检测，进而建立检测细胞内ROS的新方法。该项目研究成果可望为在细胞水平上对氧化应激事件导致的疾病的理解提供新思路，并为SERS纳米传感器的应用开发提供一定的研究基础。

活性氧物种（ROS）在生理和病理过程中发挥着重要作用，因此开发高灵敏和高选择性检测活细胞中ROS的方法对评价其生物学作用至关重要。我们基于表面增强拉曼散射技术（SERS）灵敏度高、检测时间短、水干扰小及可原位检测的优点，构筑了新型纳米复合结构，实现了活细胞内ROS的高选择性及高灵敏度检测，在细胞水平上研究了相关的基础性科学问题。创新性地研发了便携性高、价格便宜的手写型银纳米阵列，柔性棉签基底，提高了纳米结构的稳定性和重复性；利用简单的抽滤技术开发了复合薄膜基底，集快速富集和SERS检测为一体，实现了待测分子的高灵敏快速分析，解决了常规SERS基底活性低的问题。在此基础上，通过在纳米结构表面修饰芳基硼酸，基于H2O2氧化芳基硼酸释放出苯酚，构建了新型纳米传感器，实现了细胞中H2O2的高选择性检测，检出限低到80 nM。基于•OH诱导DNA链裂解，设计并构建了荧光-SERS双信号纳米传感器，对细胞中•OH实现了高选择性检测，检测限低至10 nM。进一步，通过在核-壳结构表面修饰pH响应的荧光探针分子和SERS信号分子制备了荧光-SERS双信号纳米探针，实现了细胞内pH的双信号传感，成功区分了正常细胞和癌细胞。研究成果有望为新型SERS纳米传感器的构建提供新思路，为SERS在生物分析中的应用奠定基础。