高性能石墨烯场效应晶体管生物传感器实时监测单细胞释放的一氧化氮

发布时间：2018-04-04 19:41

本文选题：生物传感器　切入点：场效应晶体管　出处：《湖北中医药大学》2017年硕士论文

【摘要】：一氧化氮既是信使分子又是神经递质,可以接收和传达信息以调节细胞活性,并且指示机体执行某些重要功能,例如神经传递、血管舒张、免疫应答和血管生成等。研究发现一氧化氮不但在动物的免疫调节、神经元通讯、平滑肌松弛等活动中充当信号分子,而且参与植物的生长发育、胁迫反应、呼吸代谢以及衰老成熟等生理过程。然而,一氧化氮作为活性气体在体内以纳摩尔浓度存在,具有相对短的半衰期(10s),且极易被氧化物或超氧化物离子氧化,使得难以精确地量化生物系统中的一氧化氮水平,因此,高效、准确、定量获取细胞及体内一氧化氮信号分子信息则是深入探索其作用机制的基础。到目前为止,许多用于一氧化氮检测的技术被应用,例如生物测定,电子磁共振光谱,鲁米诺/过氧化氢化学发光,激光诱导荧光,分光光度测定等。然而,这些方法还存在成本高、预处理复杂和灵敏度低等缺点。电化学生物传感器作为一种有效的检测工具,可以高灵敏度、实时监测一氧化氮的释放,已被广泛用于生物系统中一氧化氮的检测。然而,为实现一氧化氮检测的目的,电化学一氧化氮传感器通常需要在相对高的氧化电位(0.6V)下操作,这会干扰固有的细胞反应或诱导来自其它电化学活性分子显著的信号干扰。与安培计电极相比,场效应晶体管纳米生物传感器由于其高信噪比可实施高灵敏、实时的监测,同时,它在相对较低的检测电位(Vds=0.1V)条件下工作,这对细胞几乎没有影响。因此,使用场效应晶体管生物传感器实时检测一氧化氮具有非常重要的意义。近年来,基于场效应晶体管生物传感器用于实时监测一氧化氮的工作已经被报道。中国科学院苏州纳米所刘立伟课题组报道了基于石墨烯的场效应晶体管生物传感器检测一氧化氮气体;以色列魏茨曼科学研究院Shvarts课题组报道了一种砷化镓场效应晶体管生物传感器用于检测生理溶液中一氧化氮。然而,这些传感器的灵敏度仍然局限于微摩尔或亚微摩尔水平,且它们均未用于细胞水平的检测。最近,美国南加州大学段镶锋课题组报道了基于血红素功能化石墨烯场效应晶体管的一氧化氮传感器直接检测活细胞中释放的一氧化氮。然而,该方法未能实现单细胞水平一氧化氮的释放。细胞是生物体的基本单位,在单细胞水平的检测将增强对多种细胞过程如细胞内和细胞间通讯,细胞分化,外部刺激的生理效应和疾病状态等的认识。此外,对于诊断基于细胞的疾病,也需要开发高灵敏度的方法用于单细胞水平的检测,以获得最好的诊断能力。因此,为了实现单细胞水平一氧化氮释放的实时监测,研究高灵敏度的新型场效应晶体管生物传感器非常必要。针对以上现状和挑战,本论文立足于发展高性能场效应晶体管生物传感器以解决在单细胞水平实时监测细胞释放的一氧化氮。在本工作中,我们发展了一种基于还原氧化石墨烯和铁卟啉功能化石墨烯的场效应晶体管生物传感器,用于在单细胞水平超灵敏和高效率实时监测一氧化氮。将还原氧化石墨烯和金属铁卟啉通过π-π堆叠作用层层组装在事先制备好的石墨烯场效应晶体管生物传感器沟道表面构建卟啉功能化的石墨烯场效应晶体管生物传感器,由于石墨烯良好的导电性和卟啉分子对一氧化氮特异的催化性能,该传感器能实现对一氧化氮超灵敏和高特异性的检测。实验证明,该生物传感器展现出了优良的导电性和高催化性能,能够实时监测1pM至100nM范围内的一氧化氮,检测限可低至1pM,同时还实现了在单细胞水平超灵敏和高特异性实时监测人脐静脉内皮细胞释放的一氧化氮。因其具有微型化、超灵敏和快速响应特性,该生物传感器在生物应用和临床诊断方面有很大潜力。
[Abstract]:Nitric oxide is messenger molecule and neurotransmitter, can receive and transmit information to regulate cell activity, and indicates the body to perform some important functions, such as vasodilation, neurotransmission, immune response and angiogenesis. It is found that nitric oxide not only in animal immune, neuronal communication, act as signaling molecules of smooth muscle relaxation and other activities, and involved in plant growth and development, stress response, respiratory metabolism and senescence and other physiological processes. However, the activity of nitric oxide as a gas in the body at nanomolar concentrations, has a relatively short half-life (10s), and can easily be oxide or superoxide ion oxidation, making it difficult to accurately quantify the level of nitric oxide in biological systems, therefore efficient, accurate, quantitative, obtaining cells and in vivo nitric oxide signaling molecular information is to further explore the mechanism of the base The foundation. So far, many for nitric oxide detection technologies are applied, such as bioassay, electron paramagnetic resonance spectroscopy, hydrogen peroxide / luminol chemiluminescence, laser induced fluorescence, spectrophotometry and so on. However, these methods still have high cost, complicated pretreatment and low sensitivity. As an effective electrochemical biosensor the detection tool, can high sensitivity, real-time monitoring of the release of nitric oxide, nitric oxide detection has been widely used in biological systems. However, in order to achieve the purpose of electrochemical detection of nitric oxide, nitric oxide sensor usually in a relatively high oxidation potential (0.6V) under operation, it will interfere with the inherent cellular responses or induced from a signal the electrochemical activity of molecules significantly. Compared with the ammeter electrode, field effect transistor biosensors because of its high signal-to-noise ratio. Application of high sensitivity, real-time monitoring, at the same time, it is in a relatively low detection potential (Vds=0.1V) under the working conditions, almost no effect on the cells. Therefore, has very important significance to use field effect transistor biosensor for real-time detection of nitric oxide. In recent years, based on field effect transistor biosensor for real-time monitoring of nitric oxide the work has been reported. China Academy of Sciences Suzhou Institute for nanotechnology research group reported the Liu Liwei gas field effect transistor biosensor based on graphene oxide; Israel Weizmann Scientific Research Institute of the Shvarts research group of a GaAs FET biosensor for detection of nitric oxide in physiological solution has been reported. However, the sensitivity of these sensors is limited to micromolar or submicromolar levels, and they were not used to detect cell level. Recently, the United States and South Section Xiangfeng State University research group reported a direct detection of nitric oxide release in living cells heme functionalized graphene field effect transistor nitric oxide sensor based on single cell level. However, the release of nitric oxide. The method failed to cell is the basic unit of organism, detection at the single cell level will be enhanced to a variety of cellular processes such as cell intra - and intercellular communication, cell differentiation, understanding the physiological effects of external stimuli and disease status. In addition, the cells in the diagnosis of disease based on the method, but also need to develop a high sensitivity for the detection of single cell level, in order to obtain the best diagnostic ability. Therefore, in order to realize the real-time monitoring of single cell level of nitric oxide release. Study on the high sensitivity of the new field effect transistor biosensor is very necessary. In view of the above situation and challenges, this paper is based on the development of high The performance of field effect transistor biosensor in order to solve the problems in real-time monitoring of nitric oxide release in single cell level. In this work, we develop a reduction of graphene oxide and iron porphyrin functionalized graphene field effect transistor biosensor based on single cell level for ultra sensitive and efficient real-time monitoring of graphene oxide nitric oxide. Graphene and metal iron porphyrin reduction by tt-tt stacking layers assembled in the prepared graphene field effect transistor biosensor channel surface construction of porphyrin functionalized graphene field effect transistor biosensor, the catalytic performance of graphene with good conductivity and specific porphyrin molecules on nitric oxide, the sensor the realization of ultra sensitive and specific detection of nitric oxide. The experiment proves that the biosensor exhibits excellent electrical conductivity and high Catalytic performance, nitric oxide can real-time monitoring of 1pM to 100nM range, the detection limit can be as low as 1pM, but also achieved at the single cell level ultra sensitive and high level of real-time monitoring of human umbilical vein endothelial cell specific release of nitric oxide. Due to its miniaturization, ultra sensitive and fast response characteristics of the biosensor greatly the potential application in biological and clinical diagnosis.

【学位授予单位】：湖北中医药大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：TN386;TP212.3

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