与微流控集成的锥形纳米孔在电生命分析化学中的应用

In this project, biomimetic nanopore ( or asymmetrical nanopore-templated nanostructures) was integrated into one portable, miniaturized microfluidics device with embedded microelectrodes. This type of new micro/nano device in combination with various electroanalytical methods can be employed for fast, label-free, highly selective and sensitive detection of MicroRNA, protein biomarker,cancer cell and bacteria. Method based on ion current rectification via conically-shaped nanopore which was developed in recent years will be utilized for quantitative measurement of MicroRNA；Resistive-pulse method is another approach which can be used for quantitative target analysis via single nanopore. This method is very suitable for selective detection of protein biomarker using specific recognition agents, such as aptamer and antibody；One prominent advantage of nanopore-integrated microfludic device is the capability that allows selective fixation of cancer cell and bacteria into functionalized conical nanopore and reversible release of those analytes under certain fluidic direction, the whole process can be monitored by the change in current flowing through microelectrodes. Therefore specific detection of cancer cell and bacteria can be achieved via functionalized conical nanopore; Cancer cell-fixed nanopore facilitates further study on the interaction between cell and drug-encapsulated nanocarrier; Finally, we are going to integrate the asymmetrical nanostructures templated from conical nanopore with microfluidic device. This prepared conical-shaped nanostructure array which is integrated into device will allow easy capturing of cancer cell.The specific interaction between cancer cell and molecular recognition-functionalized nanostructures can be detected using various electrochemical methods.

本项目中，我们运用集成了仿生纳米通道(或以纳米通道为模板制作纳米结构)的微流控新的物理化学性质，并结合电分析化学技术，免标记且快速的，高选择性探测生命体系中重要的小核酸，蛋白质标记物，癌细胞和细菌。通过制作功能化的单锥形纳米孔，运用离子整流方法选择性探测小核酸(MicroRNA)；运用电阻脉冲方法探测蛋白质标记物和灭活病毒；运用锥形纳米孔不对称的纳米结构，并结合微流控在液体流动上灵活的控制能力，高选择性固定和可逆释放癌细胞核和特定细菌，整个过程可以通过集成的微电极的电流改变检测到，从而构建新的高选择性探测和控制细胞(或细菌)的微纳米芯片；选择性固定的癌细胞和细菌也可以用共聚焦显微镜来进一步研究与纳米药物胶囊的相互作用；不对称纳米孔也可以作为模板，制作功能化的不对称纳米线阵列，并被集成到微流控芯片中，在实际血样中高效的抓取和选择性探测癌细胞和细菌，整个过程可以用电化学分析方法检测。

经过20多年的发展，纳米孔分析化学取得了巨大的进步，期间发展了不同种类的纳米孔，包括蛋白质纳米孔，高分子纳米孔，玻璃纳米孔和各种无机薄膜纳米孔。于此同时，理论研究和各种功能化技术逐渐完善。研究内容从核酸测序扩展到对药物小分子，蛋白质，核酸碱基突变及其他一些重要的对象进行检测。本项目的目的就是要把纳米孔分析化学进一步发展。执行期间，为了提高纳米孔在分析化学上的应用范围和深度，把纳米孔的离子整流现象运用到分析化学。发明了光透射技术测量纳米孔孔径。借助纳米通道支撑基底，发现高分子膜材料上具备完美的离子二极管效应和离子整流现象，离子整流系数达几万倍。限域纳米孔内的立体阻碍干扰探针和探测物的相互作用，为了消除这个干扰，设计了无探针修饰的纳米孔分析平台，避免了很多技术难题。运用离子整流解释了高分子薄膜内纳米通道内羧基可以带正电，观测纳米孔内表面反应。者基于核酸探针进行了系统的探索，为纳米孔研究内容的扩展和进一步深入，提供了更多的手段和研究对象。针对固体表面核酸探针修饰，设计了新的Insertion Approach，极大地提高了信号重现性。在项目资助下，国际刊物上发表 10 篇，在纳米孔领域，占有一席之地。