金纳米粒子LSPR效应在微纳区域可逆pH传感上的应用

发布时间：2018-05-26 14:57

本文选题：i-motif + DNA　；参考：《南京大学》2017年硕士论文

【摘要】：金纳米粒子的局域表面等离子体共振(LSPR)散射效应受其自身参数及外界环境影响。单个金纳米粒子可以有效散射可见光且不发生光漂白,加上突出的生物兼容性和低细胞毒性等优势,在生物分析成像上具有极大的应用前景。金纳米团簇中粒子耦合状态改变引起的光学性质变化可以作为研究微纳区域化学过程的有力手段。pH是细胞行为的重要调节因素,细胞增殖、凋亡以及酶活性和蛋白质降解等过程都与细胞内pH变化密切相关。细胞内pH的破坏性变化可能会导致细胞器功能障碍。因此,开发生命体系中微纳尺度pH信号的获取方法对于研究细胞代谢以及获取生理病理过程关健信息意义重大。本论文旨在通过i-motif DNA与DNA的中性类似物吗啉核酸(Morpholino,MO)在低盐浓度下杂交组装得到均一、稳定的金纳米团簇,通过监测单个金纳米团簇散射光谱的变化,构建一种基于i-motif DNA分子构象变化的微纳区域pH信号分析方法。具体的实验分为两个部分:1.基于i-motif DNA与MO杂交构建Au双粒子利用i-motif DNA与MO互补杂交构建40 nm Au双粒子。将i-motif DNA和MO分别修饰到40 nm AuNPs表面,随后将两种功能化的AuNPs按照1:1的摩尔比混合组装。引入HS-PEG-COOH保护MO-Au(40),同时降低组装实验中大规模团簇产生的概率。通过优化i-motif DNA/MO分子与AuNPs的摩尔比、组装盐浓度以及加盐方式等组装条件,在低杂交效率与生成大规模团簇两个极端之间找到平衡点,得到双粒子结构为主要产物的金纳米团簇。用紫外可见光谱与透射电镜等表征手段对组装产物的结构和光学性质进行表征,并用暗场显微镜-光谱仪联用系统进一步表征单个Au双粒子的LSPR散射性质,分析其在微纳区域pH传感上的应用可行性。2.基于i-motifDNA与MO杂交构建Au core-satellite结构及pH响应性能研究利用i-motifDNA与MO互补杂交构建具有可逆pH响应的core-satellite纳米团簇。分别合成15 nm和50 nm的球形AuNPs,以DNA-Au(50)+ MO-Au(15)和 MO-Au(50)+ DNA-Au(15)两种实验方案制备 core-satellite 纳米团簇。用紫外可见光谱和透射电镜表征了两种方案的组装过程和产物结构,选择以DNA-Au(50)+ MO-Au(15)的实验方案制备的core-satellite纳米团簇为研究对象。用暗场显微镜-光谱仪联用系统表征了该core-satellite纳米团簇在微纳区域中的pH响应行为和响应可逆性,并设计控制实验验证core-satellite纳米团簇对pH可逆响应的特性来自于i-motifDNA的构象变化。
[Abstract]:The local surface plasmon resonance (LSP) scattering effect of gold nanoparticles is affected by its own parameters and the external environment. Single gold nanoparticles can effectively scatter visible light without photobleaching, combined with outstanding advantages of biological compatibility and low cytotoxicity, so it has a great application prospect in biological analysis imaging. The change of optical properties caused by the change of particle coupling state in gold nanoclusters can be used as a powerful means to study the chemical process of micro-nano region. Ph is an important regulator of cell behavior and cell proliferation. Apoptosis, enzyme activity and protein degradation are closely related to intracellular pH changes. Destructive changes in intracellular pH may lead to organelle dysfunction. Therefore, it is of great significance to develop the method of obtaining micro- and nanoscale pH signal in life system for studying cell metabolism and obtaining key information of physiological and pathological processes. The aim of this thesis is to obtain uniform and stable gold nanoclusters by hybridization of i-motif DNA and morpholine nucleic acid (MOO), a neutral analogue of DNA, at low salt concentration, and to monitor the variation of scattering spectra of single gold nanoclusters. A method of pH signal analysis in micro and nano region based on conformation change of i-motif DNA molecule was constructed. The specific experiment is divided into two parts: 1. Two au particles were constructed based on i-motif DNA and MO hybridization. The 40 nm au double particles were constructed by i-motif DNA and MO complementary hybridization. I-motif DNA and MO were modified to 40 nm AuNPs surface, and then the two functionalized AuNPs were assembled at 1:1 molar ratio. HS-PEG-COOH is introduced to protect MO-Au-40, and the probability of large scale cluster formation in assembly experiment is reduced at the same time. By optimizing the molar ratio of i-motif DNA/MO molecule to AuNPs, the concentration of salt and the method of salt addition, the equilibrium point was found between the low hybridization efficiency and the formation of large clusters, and the gold nanoclusters with two-particle structure as the main products were obtained. The structure and optical properties of the assembled products were characterized by UV-Vis spectroscopy and transmission electron microscopy. The LSPR scattering properties of single au double particles were further characterized by dark field microscopy and spectrometer. The feasibility of its application in micro-nano region pH sensing was analyzed. Structure and pH response of au core-satellite based on i-motifDNA and MO Hybridization; core-satellite nanoclusters with reversible pH response were constructed by complementary hybridization of i-motifDNA and MO. Spherical AuNPs at 15 nm and 50 nm were synthesized respectively. Core-satellite nanoclusters were prepared by DNA-Au-50) MO-Au-15) and MO-Au-50) DNA-Au-15). The assembly process and product structure of the two schemes were characterized by UV-Vis spectroscopy and transmission electron microscopy. The core-satellite nanoclusters prepared by DNA-Au-50) MO-Au-15) were selected as the object of study. The pH response behavior and reversibility of the core-satellite nanoclusters in micro and nano region were characterized by a dark field microscope and spectrometer system. The control experiments were designed to verify that the reversible response of core-satellite nanoclusters to pH was due to the conformation change of i-motifDNA.
【学位授予单位】：南京大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：O614.123;TB383.1

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