功能纳米材料在禽白血病病毒生化分析中的应用

发布时间：2018-02-28 07:15

本文关键词： 电化学免疫传感器石墨烯量子点包铜去铁蛋白磁性碳纳米管中空介孔二氧化硅　出处：《山东农业大学》2015年硕士论文　论文类型：学位论文

【摘要】：禽白血病是由不同的RNA逆转录病毒引起的可传播性的家禽肿瘤性疾病。近几年来,由禽白血病引起的家禽死亡及淘汰率逐年上升,造成的经济损失逐年增加。除此之外,由禽白血病感染之后造成的免疫抑制和对疫苗的应答能力下降等间接损失更是无法估量。本文研究了石墨烯量子点、包铜去铁蛋白、磁性碳纳米管和中空介孔二氧化硅等纳米材料及其复合材料的电化学性能,并结合分子识别、直接氧化还原、纳米材料的类酶催化等方法,构建了几种不同类型的电化学免疫传感器,实现了对禽白血病病毒的超灵敏检测。主要分为以下三部分:(1)本研究用石墨烯量子点和包铜去铁蛋白作为信号放大物质,制作了一种新颖的检测禽白血病病毒的电化学免疫传感器。分别以石墨烯量子点修饰电极连接一抗,复合四氧化三铁磁珠连接二抗,实现三明治式的电极组装。然后用包铜去铁蛋白结合Fe3O4@GQDs纳米粒子,将电信号活性物质-铜纳米粒子引入了传感器体系中。当改变环境的pH值,电信号物质被释放到溶液中,可以通过微分脉冲伏安法对释放出来的电信号物质进行检测。由于石墨烯量子点有巨大的比表面积和大量的含氧官能团,可以结合更多的抗体,从而放大体系的电化学信号。去铁蛋白提供了容纳信号物质的巨大空腔,使得检测信号进一步被放大。实验中,通过对比不同标记的信号探针,证实了传感器的信号放大作用。禽白血病病毒浓度在102.08到104.50 TCID50/mL范围内呈很好的线性关系,在信噪比为3时,检测限为115 TCID50/mL。实验结果显示该传感器具有高的灵敏度,好的重复性和稳定性。(2)本研究首次以中空介孔二氧化硅和去铁蛋白联合构建了一种受pH控制的新型免疫传感器,实现了对禽白血病病毒的高灵敏检测。首先,制备了环糊精功能化的石墨烯,中空介孔二氧化硅、包铜去铁蛋白和二抗的复合物(HMS-apoferritin-Cu/Ab2-BSA),利用TEM对CD-GS、HMS和HMS-apoferritin-Cu/Ab2-BSA探针进行了表征。然后,以CD-GS作为基底、HMS-apoferritin-Cu/Ab2-BSA作为探针构建电化学免疫传感器。利用CV对修饰电极进行了表征,研究了CD-GS的电化学性能,探讨了富集时间和富集电位对响应信号的影响。最后利用DPV实现了对ALVs-J的高灵敏检测,并探讨了免疫传感器的选择性、重现性和稳定性。(3)基于一种多重还原银的信号放大方法,构建了一种三明治式的电化学免疫传感器,实现了对禽白血病病毒的检测。首先,用CD-GS修饰的裸电极连接一抗,然后用牛血清白蛋白进行活性位点的封闭,以防止非特异性的吸附;经过三明治式的电极组装,CIMC-Au-HAs/Ab2-BSA探针被引入到免疫传感器中,再把组装好的电极放入含银离子的溶液中进行银的还原,还原出来的银通过微分脉冲伏安法进行检测。为了说明磁性碳纳米管优越的性能,纳米金的催化还原能力和腐殖酸的优良吸附能力,我们对不同的纳米探针进行了比较。禽白血病病毒浓度在102.05到104.50 TCID50/mL范围内呈很好的线性关系,在信噪比为3时,检测限为110 TCID50/mL。实验结果显示该传感器具有高的灵敏度,潜在的临床应用能力。
[Abstract]:Avian leukosis is caused by different RNA retrovirus can spread the avian tumor diseases. In recent years, caused by avian leukosis poultry deaths and elimination rate increased year by year, the economic losses caused by increased year by year. In addition, caused by avian leukosis virus infection after immune suppression and vaccine response ability the decline of indirect losses is incalculable. This paper studied graphene quantum dots, copper coated iron protein, the electrochemical properties of carbon nanotubes and hollow mesoporous silica nanoparticles and their composite materials, and combined with molecular recognition, direct redox, nano materials enzymatic method, constructed electrochemical immunoassay several different types of sensors, to achieve the ultra sensitive detection of avian leukosis virus. Mainly divided into the following three parts: (1) this study used as graphene quantum dots and copper coated iron protein The signal amplification material, electrochemical immunosensor fabricated a novel avian leukosis virus detection. With graphene quantum dot modified electrode is connected with a resistance, connecting the two anti magnetic composite ferroferric oxide electrode, realize the sandwich assembly. Then use the copper to iron binding protein of Fe3O4@GQDs nanoparticles, the electrical signal of active substances - copper nanoparticles the introduction of the sensor system. When changing the pH value of the environment, physical signals are released into the solution, by differential pulse voltammetry to detect signal substances released. Because graphene quantum dots have huge surface area and a large number of oxygen-containing functional groups, can be combined with more antibodies, thereby amplifying the electrochemical signal system. To provide a huge iron protein cavity containing signal substances, so the detection signal is further amplified. In the experiment, by comparing with standard Remember that a signal probe, the sensor signal amplification. Avian leukosis virus concentration showed a good linear relationship in the range of TCID50/mL 102.08 to 104.50, when the signal-to-noise ratio is 3, the detection limit was 115 TCID50/mL.. The experimental results show that the sensor has high sensitivity, good repeatability and stability. (2) this is the first study to hollow mesoporous silica and iron protein and constructed a new immune sensor controlled by pH, the realization of avian leukosis virus sensitive detection. Firstly, cyclodextrin functionalized graphene prepared hollow mesoporous silica, copper clad iron protein and complex two - the use of CD-GS (HMS-apoferritin-Cu/Ab2-BSA), TEM, HMS and HMS-apoferritin-Cu/Ab2-BSA probe were investigated. Then, using CD-GS as substrate, HMS-apoferritin-Cu/Ab2-BSA as a probe to construct electrochemical immunosensor. The modified electrode was characterized by CV, the electrochemical properties of CD-GS were investigated and discussed the enrichment time and accumulation potential of the response signal. Finally realizes the high sensitive detection of ALVs-J by DPV, and discusses the selective immune sensor, reproducibility and stability. (3) a multiple signal reduction of silver amplification method based on the construction of a sandwich type electrochemical immunosensor, realized the detection of avian leukosis virus. First of all, the bare electrode modified with CD-GS connection of a resistance, then the active site of bovine serum albumin closed, to prevent nonspecific adsorption; after the electrode sandwich assembly, CIMC-Au-HAs/Ab2-BSA probe is introduced to the immune sensor, then restore the assembled electrode into the solution containing silver ion in silver, the reduction of silver by differential pulse voltammetry for detection. The magnetic properties of carbon nanotubes superior catalytic reduction of gold nanoparticles, excellent adsorption ability and humic acid, we compare the different nano probe. Avian leukosis virus concentration showed a good linear relationship in the range of TCID50/mL 102.05 to 104.50, when the signal-to-noise ratio is 3, the detection limit is 110 TCID50/mL. the experimental results show that the sensor has high sensitivity, clinical application potential.

【学位授予单位】：山东农业大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S852.65;TB383.1

【共引文献】