基于超微电极的快速扫描循环伏安法研究纳米材料的神经毒性

Because nanomaterials can cause a series of changes in cell morphology and function, they pose potential risk to humans due to their unique physiochemical properties.The nerve cells are non-renewable, so the damage to nerve cells has very huge effect on the organisms. At present, fluorescent dyes were used in the most detecting methods for neurotoxicity of naomaterials. However, the detection would be interfered by nanomaterials due to their high surface activity. This project aims to detect the neurotoxicity of naomaterials by the change in the release of dopamine and NO or more species before and after the exposure of nerve cells to nanomaterials at ultramicroelectrodes with amperometry and fast-scan cyclic voltammetry methods. Furthermore, microelectrodes array can also be used to detect several cells simultaneously and assess the neurotoxicity of nanomaterials with the information on the interaction between cells. Due to the small size of ultramicroelectrodes, they will not destroy the biological tissue and can be used in micro-analysis and in vivo analysis by inserting them into organisms. Due to the high sensitivity, high selectivity and good spatial and temporal resolution, amperometry and fast-scan cyclic voltammetry are the most useful tools for detection of transmitter releasing from single nerve cell. The project is expected to establish an accurate and rapid method to assess the neurotoxicity of nanomaterials by detection of neurotransmitters releasing from nerve cells. This method also shows high sensitivity, good selectivity, high temporal and spatial resolution.

纳米材料因可引起细胞一系列形态和功能变化而对生命体的健康产生着负面影响。因神经细胞的不可再生性，神经细胞的损伤对生物体的影响更加巨大。目前检测纳米材料神经毒性的方法中多使用荧光染料，由于纳米材料高的表面活性，可以吸附染料分子从而对检测结果产生干扰。本项目拟在超微电极上以安培法和快速扫描循环伏安法测定暴露于纳米材料前后神经细胞受激时释放多巴胺、一氧化氮等物质的变化情况，以多指标准确评价纳米材料的神经毒性。进而采用微电极阵列同时检测多个细胞，通过细胞之间信号转导的变化为纳米材料的神经毒性提供更进一步的评价。由于超微电极半径极小，不会破坏生物组织，可插入生物体内进行微区分析和活体分析。拟用的安培法和快速扫描循环伏安法由于具有高灵敏度、高选择性和高时空分辨率等优点，是检测单细胞中神经递质释放的最有用工具。故本项目有望建立一种高灵敏度、高选择性、高时空分辨且准确、快速检测纳米材料神经毒性平台。

本项目拟利用超微电极测定暴露于纳米材料前后神经细胞受激时所释放多巴胺、一氧化氮等活性物质的量的变化，实现纳米材料对神经细胞毒性的多指标准确评价。但由于单细胞中生物活性物质的含量极低，而且活体中物质种类繁多，所以提高方法的灵敏度和选择性显得很重要；此外，细胞的个体差异性及细胞中小分子代谢物的含量处在动态平衡之中，所以为了反映单细胞中活性物质的真实状态和变化，需建立单细胞水平的研究及细胞中活性物质的快速测定，需实时快速检测活性物质。基于此两点，本项目在提升测定灵敏度、选择性和响应速度上进行了相关研究。通过电极表面修饰疏水单分子层及通过热处理使碳纤维电极表面形成疏水性多孔结构，改变了电化学发光微环境，增强了电化学发光测定的灵敏度；通过在不同尺寸超微电极表面电沉积形成金纳米花，增大了电极活性面积、减小了空间位阻，使所构建的金纳米花修饰超微电极具有较高的灵敏度和较快的响应速度；在二氧化硅纳米粒子中引入富含氨基的PEI，基于金属离子与氨基强的配位作用，使二氧化硅纳米粒子中掺入高浓度的金属离子，所以以金属离子作为电活性物质的电化学分析方法的灵敏度得以提高；此外，基于纳米粒子与超微电极之间的相互作用，利用电化学发光法实现了单一硬质纳米粒子表面状态的表征。在此基础上，基于单细胞与超微电极之间的相互作用，实现单细胞的选择性测定。