基于纳米孔三明治阵列的肿瘤标志物定量检测研究

Tumor markers detection technique based on nanopores array sensor has become an important potential method for the early stage cancer diagnosis because of the rapid response capabilities. At present, quantitative detection technique of tumor markers based on nanopores array sensor exists disadvantages of low fabrication efficiency and inaccurate position of modified antibodies. The silicon nitride/silicon/silicon nitride (Si3N4/Si/Si3N4) nanopores sandwich arrays will be proposed, which is used to control the position of modified antibodies precisely. In addition, the detection of multiple tumor markers in a parallel way will be also studied in this research program. The relationship between the parameters of AAO template and the size of silver nanoparticles and their distribution on the surface of silicon wafers will be established to reveal the silver nanoparticle-assisted HF/H2O2 etching silicon nanopores mechanism. Then, rapid fabrication of Si3N4/Si/Si3N4 nanopores sandwich structure technology is obtained in controllable way. In theory, the intrinsic relationship between nanopores sandwich structure’s blockade ionic current and tumor markers will be established based on the molecular dynamics (MD) simulations. The principle of tumor markers’ concentration detection and the controllability of modified position of antibody molecules will be also established. Based on the difference binding force between antibody molecules and materials in nanopores sandwich can be well controlled. The theory and method of quantitative detection of tumor markers will be built. The parallel detection method for multiple tumor markers will be proposed after three-channel nanopores sandwich arrays sensor is fabricated and studied. This project promotes the theory and application development of micro-nano controllable manufacturing, and provides accurate detection methods for the early stage cancer diagnosis.

具有快速响应的特点纳米孔阵列传感器已成为一项极具竞争力的肿瘤标志物检测手段，是癌症早期诊断的重要研究方法。目前，基于纳米孔阵列的肿瘤标志物定量检测存在抗体修饰位置不准与可控制造效率低问题。本课题提出氮化硅/硅/氮化硅纳米孔三明治阵列实现肿瘤标志物抗体修饰位置的主动控制，实现对多种肿瘤标志物并行检测研究。建立AAO模板特征参数与银纳米颗粒尺寸及其在硅片表面分布关系，揭示银纳米颗粒辅助HF/H2O2刻蚀硅纳米孔机理，获得Si3N4/Si/Si3N4纳米孔三明治阵列快速可控制造方法；通过分子动力学模拟，阐明纳米孔三明治结构阻塞电流与肿瘤标志物量的关联机制，弄清抗体分子修饰位置主动控制与肿瘤标志物精准定量检测的机制，建立肿瘤标志物定量检测理论与方法；实现三通道肿瘤标志物并行检测纳米孔三明治传感器制造；提出多种肿瘤标志物并行检测方法；推动微纳可控制造理论与应用发展，为癌症早期诊断提供精准的检测手段。

针对纳米孔阵列的肿瘤标志物定量检测存在抗体修饰位置不准与可控制造效率低问题，本项目开展纳米孔阵列的肿瘤标志物定量检测研究，研究内容包括纳米孔阵列可控制造技术、纳米孔三明治结构的肿瘤标志物定量检测理论和肿瘤标志物检测。通过采用Al2O3/Au/Si3N4纳米孔三明治实现肿瘤标志物抗体修饰位置的主动控制，实现了对肝癌标志物癌胚抗原（AFP）的双通道并行检测研究。建立了AAO 模板特征参数与银纳米颗粒尺寸及其在硅片表面分布关系、电子束和离子束与纳米孔直径的关系，揭示了银纳米颗粒辅助 HF/H2O2刻蚀硅纳米孔的无氧化硅参与及光照条件下氧化硅参与的刻蚀机理、电子束收缩机理和离子束刻蚀机理，获得了氮化硅、石墨烯、二硫化钼以及Si3N4/Si/ Si3N4和Al2O3/Au/Si3N4 纳米孔三明治阵列结构快速可控制造方法；通过COMSOL模拟，建立了纳米孔直径大小、纳米孔形状、电解质浓度等因素对阻塞电流的影响机制，建立了纳米孔三明治结构的肿瘤标志物定量检测理论与方法；实现了双通道阵列纳米孔肝癌标志物（AFP）并行检测，单孔、2*2和2*3阵列纳米孔最小检测灵敏度为70fM。同时本项目还补充了氮化硅阵列纳米孔、石墨烯纳米孔检测牛血清蛋白不同的过孔姿态，以及氮化硅纳米孔实现AuNPs-DNAs不同易位形式的区分；本项目推动了微纳可控制造理论与应用的发展，为疾病早期诊断提供了精准的检测手段。项目发表论文5篇（其中SCI 论文2篇），发明专利授权8项。培养研究生6名，其中取得硕士学位4名，在读2名。项目经费共计31.06万元，支出26.6671万元，各项支出与经费预算基本相符，剩余经费计划用于本项目研究后续支出。