多功能化双金属纳米材料的制备及其在电致化学发光免疫分析上的应用

发布时间：2018-06-19 05:10

本文选题：电致化学发光 + 功能化双金属纳米材料　；参考：《西南大学》2017年硕士论文

【摘要】：随着纳米技术的迅速发展,基于金属纳米材料的信号放大方法对提高免疫传感器的灵敏度和选择性表现出巨大的潜力。其中双金属纳米材料在电催化活性等方面比相应的单金属拥有更优越的性质。但目前如何赋予双金属纳米材料更多的功能,从而简化实验操作、提高作用效率仍然存在挑战。本论文的工作主要集中在制备多种功能性双金属纳米复合材料、发光试剂功能化壳核纳米材料,结合电致化学发光(ECL)传感技术和生物技术,构建新型、高灵敏的免疫传感器。成功合成了共反应试剂功能化的六八面体壳核材料、异鲁米诺功能化多面体状壳核纳米粒子和自催化型鲁米诺衍生物功能化的钯铜纳米立方@碳纳米角复合物,也提出了共反应试剂或催化剂对发光物可能的作用机理。基于上述多功能化纳米复合材料,发展了一系列新型ECL生物分析新方法,实现了对多种蛋白质的灵敏检测。本论文的研究工作主要分为以下几部分:1.基于共反应试剂功能化的六八面体壳核纳米材料构建电致化学发光免疫传感器将共反应试剂置于检测底液中不但会增加试剂用量,而且也会带来测量误差。本研究利用六八面体壳核纳米材料作为载体成功实现了共反应试剂的大量固载以及ECL的信号放大,在简化实验操作的同时也提高了传感器检测的灵敏度。研究证明当测试底液中有诺氟沙星(NFLX)存在时S2O82-能产生非常强的ECL信号,强度约是S2O82-单独存在时的350倍。为了实现NFLX在电极表面的的大量固载,利用聚酰胺树枝状高分子的大量的端氨基,通过酰胺键将NFLX修饰在聚酰胺聚合物(PAMAM)表面从而形成PAMAM-NFLX复合物。该复合物作为新颖的共反应试剂能有效放大S2O82--O2体系的ECL信号。同时,合成了凸状的壳核Pd@Au六八面体纳米颗粒(Pd@AuHOHs),该粒子不仅拥有良好的生物相容性和电子传导能力等优点,通过促进S2O82--O2体系ECL反应过程中中间自由基的产生,Pd@AuHOHs还能进一步提高ECL信号。此外,由于具有大的比表面积,Pd@AuHOHs作为纳米载体能用来大量固载第二抗体以及PAMAM-NFLX复合物。基于此,构建夹心型ECL免疫传感器用于检测促甲状腺激素(TSH)。该传感器具有高的灵敏度和特异性,成功地实现了对临床血清样本中TSH的检测。2.基于发光功能化壳核纳米颗粒和二茂铁衍生物的电致化学发光免疫传感器的研究传统的直接合成鲁米诺或其衍生物功能化纳米材料的方法得到的发光功能化纳米材料常为单金属且大多需要高温煮沸的条件。研究表明双金属纳米材料在电催化活性等方面比相应的单金属拥有更优越的性质。鉴于此,本研究采用鲁米诺的一种衍生物N-4-氨基丁基-N-乙基异鲁米诺(ABEI)作为还原剂和发光试剂在常温下成功合成了ABEI功能化的Pd@Au壳核纳米颗粒(ABEI-Pd@AuNPs)。该法成功实现了ABEI分子的固载,这样就避免了后期修饰。此外,由于双金属纳米颗粒对H2O2良好的催化性质,ABEI-Pd@AuNPs能够增强ABEI-H2O2体系的ECL信号。为了进一步提高发光效率,利用L-半胱氨酸将催化剂羧基二茂铁(Fc)连接在ABEI-Pd@AuNPs的表面。ABEI和Fc同时修饰在Pd@Au纳米颗粒上意味着在纳米颗粒表面二者的实际浓度远高于将它们置于溶液中,从而能更好地实现催化。将该双功能化的Pd@Au壳核纳米颗粒作为载体固载第二抗体,利用夹心法构建了信号增强型的ECL免疫传感器,实现了对糖尿病肾病潜在标志物IV-型胶原(Col IV)的检测。最终该传感器达到了1 pg·mL-1到10 ng·mL-1的宽检测范围。该工作提供了一种新颖的信号放大方法,也拓宽了ABEI-H2O2体系在生物分析中的应用。3.自催化型发光体与PdCu@碳纳米角杂化物构建电致化学发光免疫传感器通常基于鲁米诺(luminol)的ECL反应中,催化剂对luminol的信号放大是通过分子间的相互作用实现的。然而这种分子间的作用通常伴有较大的能量损失,限制了luminol与其催化剂之间的作用效率。为了实现将发光物luminol固载的同时提高其与催化剂之间的作用效率,本研究首次提出了基于自催化型发光衍生物和碳纳米角杂化物放大luminol/H2O2体系ECL信号的放大策略,并成功构建了用于检测心衰标志物的超灵敏ECL免疫传感器。将3,4,9,10-傒四甲酸(PTCA)与luminol偶联起来形成自催化的发光体PTC-Lu,由于PTCA强的增强效果,该发光体具有极好的ECL性质;且由于芳香性,使得PTC-Lu更容易实现固载。通过π-π堆积作用将该自催化发光体组装在PdCu@SWCNHs纳米杂化物上,得到PTC-Lu功能化的PdCu@SWCNHs(PTC-Lu/PdCu@SWCNHs)。通过这种方式明显提高了SWCNHs的水溶性和稳定性。同时,PdCu@SWCNHs杂化物作为模拟酶对H2O2展现出电催化性质,因此能进一步放大luminol/H2O2体系的ECL信号。此外,由于PdCu纳米立方的高比表面积和良好的生物相容性,能够实现对二抗蛋白的大量固载。基于此,构建了信号增强型的免疫传感器,并成功实现了对临床人体血清样本中N-末端B型钠尿肽(NT-proBNP)的检测。本工作在解决luminol固载问题的同时,在提高催化剂作用效率方面也得到突破。此外,该发光功能化纳米材料的合成不仅改善了SWCNHs的水溶性和稳定性,其中PdCu@SWCNHs也展示出良好的催化活性,该新颖的纳米杂化物作为模拟酶在生物技术和临床诊断有着潜在应用价值。
[Abstract]:With the rapid development of nanotechnology, the signal amplification based on metal nanomaterials has shown great potential to improve the sensitivity and selectivity of the immunosensor. In which the electrocatalytic activity of bimetallic nanomaterials is more superior than that of the corresponding monometallic metal. The work of this paper is mainly focused on the preparation of functional bimetal nanocomposites, luminescent reagent functionalized shell nanomaterials, electrochemiluminescence (ECL) sensing technology and biological technology, to construct a new and highly sensitive immune sensor. The functionalized 68 hedral shell material, the functionalized polyhedron shell nanoparticles and the autocatalytic luminol derivative functionalized palladium copper nanomaterials, also proposed the possible mechanism of the co reaction reagent or catalyst on the luminescence. Based on the above functionalization, the function of the co reacting reagent was synthesized. Nanocomposites have developed a new series of new methods for ECL bioanalysis to achieve sensitive detection of various proteins. The research work of this paper is divided into the following parts: 1. an electrochemically immunized sensor based on the functionalized 68 surface shell nanomaterials of CO reacting reagents for the detection of CO reagents In this study, the dosage of reagents will not only increase the amount of reagents, but also the measurement error. In this study, a large number of solid loading of the co reagents and the signal amplification of ECL were successfully realized by using the 68 surface shell core nanomaterials as a carrier. The sensitivity of the sensor was also improved while the experimental operation was simplified. When norfloxacin (NFLX) exists, S2O82- can produce a very strong ECL signal, which is about 350 times as strong as S2O82- alone. In order to achieve a large amount of immobilization of NFLX on the surface of the electrode, a large number of amino terminated amino groups of polyamide dendrimers are used to modify the NFLX on the surface of polyamide polymer (PAMAM) through the amide bond to form a PAMAM-NFLX complex. The compound can effectively amplify the ECL signal of the S2O82--O2 system as a novel co reacting reagent. At the same time, the protruded shell of the shell Pd@Au 68 surface nanoparticle (Pd@AuHOHs) has been synthesized. The particle not only has the advantages of good biocompatibility and electron conduction, but also through the intermediate freedom to promote the ECL reaction in the S2O82--O2 system. The Pd@AuHOHs can further improve the ECL signal. In addition, because of the large specific surface area, Pd@AuHOHs can be used as a nanoscale to immobilizing second antibodies and PAMAM-NFLX complexes. Based on this, the sandwich ECL immune sensor is constructed for the detection of thyroid stimulating hormone (TSH). The sensor has high sensitivity and specificity. A successful implementation of the detection of TSH in clinical serum samples.2. based on electrochemiluminescent immunosensor based on luminescent functionalized putamen nanoparticles and two ferrocene derivatives, the traditional direct synthesis of Lumino or its derivative functionalized nanomaterials is usually single metal and large In this study, a derivative of Lumino's N-4- amino butyl -N- ethyl ISO Lumino (ABEI) was used as a reducing agent and luminescent reagent to synthesize ABEI function successfully at normal temperature. The modified Pd@Au shell nanoparticles (ABEI-Pd@AuNPs). This method successfully realized the immobilization of ABEI molecules, thus avoiding the later modification. In addition, because of the good catalytic properties of the bimetal nanoparticles on H2O2, ABEI-Pd@AuNPs can enhance the ECL signal of the ABEI-H2O2 system. In order to improve the luminous efficiency, L- cysteine will be used as the catalyst. Carboxyl two ferrocene (Fc) is connected to the surface of the ABEI-Pd@AuNPs on the surface.ABEI and Fc simultaneously on the Pd@Au nanoparticles, which means that the actual concentration of the two of the nanoparticles is much higher than that in the solution and thus can be better catalyzed. The dual functionalized Pd@Au shell nanoparticles are used as the carrier to immobilizing second antibodies and using the sandwich. A signal enhanced ECL immunosensor was constructed and the detection of IV- type collagen (Col IV) of the potential marker of diabetic nephropathy was realized. The sensor finally reached a wide detection range of 1 pg. ML-1 to 10 ng. ML-1. This work provides a novel signal amplification method and also widens the application of.3 in the biological analysis of.3. The electrochemiluminescence immunosensor of the autocatalytic luminescence and the PdCu@ carbon nanohybrids is usually based on the ECL reaction of Lumino (luminol), and the signal amplification of the catalyst to luminol is realized by intermolecular interaction. However, the interaction of these molecules is usually accompanied by greater energy loss, limiting the luminol and its effect. In order to improve the efficiency between the luminol and the catalyst, the amplification strategy based on the autocatalytic luminescent derivatives and carbon nano corner hybrids to amplify the ECL signal of the luminol/H2O2 system was first proposed, and a successful construction of the markers for the detection of heart failure was successfully constructed. The ultra sensitive ECL immunosensor is coupled with 3,4,9,10- four formic acid (PTCA) and luminol to form a self catalyzed luminescent body PTC-Lu. Due to the strong enhancement effect of PTCA, the luminescent body has excellent ECL properties. And because of its aromatic property, the PTC-Lu is more easily immobilized. The autocatalytic luminescent body is assembled in PdCu@SWCNH by pion pion accumulation. The PTC-Lu functionalized PdCu@SWCNHs (PTC-Lu/PdCu@SWCNHs) was obtained on the s nanohybrids. By this way, the solubility and stability of SWCNHs was obviously improved. At the same time, PdCu@SWCNHs hybrids were used as analog enzymes to exhibit the electrocatalytic properties of H2O2, thus further amplifying ECL signals of the luminol /H2O2 system. In addition, PdCu nanometers were used. The high specific surface area and good biocompatibility can achieve a large amount of immobilization of two anti protein. Based on this, a signal enhanced immunosensor is constructed and the detection of B type natriuretic peptide (NT-proBNP) of the N- terminal in the serum samples of the clinical human body is successfully realized. This work improves the catalyst at the same time to improve the luminol immobilization problem and improve the catalyst. In addition, the synthesis of the luminescent functionalized nanomaterials not only improves the water solubility and stability of SWCNHs, but also PdCu@SWCNHs shows good catalytic activity. The novel nano hybrid as a mimic enzyme has potential application value in biological technology and clinical diagnosis.
【学位授予单位】：西南大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：TB383.1;TP212

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