基于电化学分子整流器信号放大技术的恶性肿瘤microRNAs检测方法研究

It is of great significance to develop novel miRNAs detection method for disease diagnosis, drug treatment and prognosis of tumor patients. Recently, electrochemical biosensors have attracted much attention in miRNAs detection, however, the signal amplification issue is always a bottleneck for their further development. Here we propose to adopt the novel electrochemical current rectification (ECR) strategy via specific nucleic molecular recognition to enhance miRNAs detection with high sensitivity and selectivity. In a typical ECR design, the redox-tags on surface after miRNAs binding will be continuously replenished by solution-phase redox molecules, in which an unidirectional electron transfer arises that efficiently amplifies the current response. Furthermore, the biosensor will be transferred on solid interdigitated array electrodes, where signal amplification will be improved through redox cycle between the neighboring electrodes. Special three-way junction DNA will be designed by introducing locked nucleic acid monomers to improve the selectivity between targets and their similar sequences, including mismatched miRNAs, precursor miRNAs and the family RNA members. In addition, the miRNAs in clinical patients' blood and in total RNAs extracted from tumoral and normal cells will be compared, as well as the expression levels after drugs treatment to monitor the anticancer efficiency. This work will improve the development of electrochemical biosensor in the field of miRNAs detection, and provide guidance for novel molecular current rectifier in signal amplification and disease diagnosis.

发展恶性肿瘤miRNAs检测的新方法对疾病诊断、药物治疗及预后判断意义重大。近年来，电化学生物传感器在miRNAs检测中备受关注,但如何实现信号放大成为制约其发展的瓶颈问题。本项目拟利用核酸结构的多样性，在表面构筑基于分子整流器效应的电化学器件实现高灵敏度和特异性检测。通过溶液相分子持续补充由表面miRNAs结合后引起的电荷传递，进行单向可控的信号增强，并将其迁移到微间距叉指阵列电极芯片表面，进一步放大信号。实验将引入含锁核酸碱基的特殊“三向连接”结构，增强对目标miRNAs与其错配序列、前体miRNA及同源家族序列间的选择性。此外，基于分子整流器信号放大的电化学芯片将应用在临床患者血液分析，对比肿瘤细胞和正常细胞内表达水平，并针对典型抗癌药物进行药物疗效预后监测。本项目将推动电化学传感器在肿瘤miRNAs检测方面的发展, 探索分子整流器的信号放大功能及应用，具有重要的科学意义和实用价值。

本项目构建了电化学生物传感器对不同生物标记物分子进行检测，包括恶性肿瘤miRNAs等，所设计的新构象、新方法对疾病诊断、药物治疗及预后判断意义重大。利用了核酸结构的多样性，在表面构筑基于分子整流器效应的电化学器件实现高灵敏度和特异性检测。通过溶液相分子持续补充由表面miRNAs结合后引起的电荷传递，进行单向可控的信号增强。引入含锁核酸碱基的特殊“三向连接”结构，增强对目标miRNAs与其错配序列、前体miRNA及同源家族序列间的选择性。此外，将应用在临床患者血液分析，对比肿瘤细胞和正常细胞内表达水平，并针对典型抗癌药物进行药物疗效预后监测。同时拓展不同核酸构象的改变对前列腺癌标记物和神经递质类分子的检测，并对比其在不同生理条件下的表达水平等。项目的执行能够推动电化学生物传感器在肿瘤及相关疾病标记物方面的探索和应用。