【作者】 刘松；

【导师】 赵元弟；

【作者基本信息】 华中科技大学 ， 生物医学工程， 2006， 硕士

【摘要】 硅纳米颗粒作为一种新型的纳米材料,在生物领域的应用近来受到越来越多的关注,并获得较大的发展。其中,基于荧光硅纳米颗粒的超灵敏的生物成像进展最快,同时也是研究者最感兴趣的领域。荧光硅纳米颗粒的制备有两种方法:St?ber法和微乳液法。其中微乳液法通过改变其反胶束的大小可控制硅颗粒的粒径大小,而且制备的硅颗粒具有较好的均一性,因而受到更多的青睐。采用微乳液法可在硅颗粒中包入水溶性荧光染料以及量子点,疏水性的有机荧光染料在经过改性后也可包入到硅颗粒的核中。与普通的荧光染料相比,硅纳米颗粒的荧光强度和荧光稳定性显著增强。硅纳米颗粒的表面可通过物理吸附或共价结合与不同的生物分子如DNA、抗体结合,从而拓展其在生物分析或生物成像领域的应用。本文首先采用“一步法”合成了表面含氨基的硅纳米颗粒,即通过正硅酸乙酯(TEOS ,水解形成硅颗粒)和氨基硅烷γ-Aminopropyltrimethoxysilane(APS)同时水解并在硅颗粒表面共聚合而修饰上氨基。继而采用“两步法”向硅纳米颗粒表面引入氨基,即在硅纳米颗粒的合成完成后通过氨基硅烷如APS等在硅颗粒表面水解缩合而实现硅纳米颗粒的表面氨基修饰。通过比较两种方法所得硅纳米颗粒的荧光强度、粒度分布以及单分散性,发现“两步法”所得纳米颗粒较佳。然后分别采用戊二醛和EDC(1-ethyl-3-3(3-dimethylaminopropyl)carbodiimide hydrochloride)、NHS(N-hydroxy-succinimide)等作为硅纳米颗粒与生物识别分子的偶连剂。戊二醛可与氨基和生物分子形成schiff碱基从而实现氨基和识别分子的偶联。而将氨基羧化衍生后,利用EDC和NHS也可与含氨基的识别分子形成稳定的酰胺键键合到硅颗粒表面。细胞实验表明,EDC和NHS的偶联效果较好。这一工作为荧光硅纳米颗粒的具体应用提供了很好的支持。更多还原

【Abstract】 As a newly developed nano-material, silica nanoparticles attracted the researchers’focus on their diversified application in biological research. Among the diverse research studies that use dye-doped silica nanoparticles, ultrasensitive bioanalysis and cellular imaging have made the most remarkable progress and attracted the greatest interest. There are two general routes for the preparation of luminescent silica nanoparticles: St?ber method and reverse micro-emulsion strategy. Reverse micro-emulsion strategy is more preferable for its convenience to control the size and uniformity of the silica nanoparticles by changing the size of the reverse micelles.A series of luminescent silica nanoparticles have been fabricated through doping various hydrophilic luminophor and quantum dot inside the core of the nanoparticles. Hydrophobic dye doped silica nanoparticle could also be attained after making these molecules hydrophilic. These silica nanoparticles are extremely bright because of the encapsulation of thousands of fluorescent dye molecules inside the silica shell. Compared with the organic luminophor, the silica nanoparticles have greater fluorescent intensity and become more photostable. Bio-molecules such as DNA、antibody could be immobilized onto the nanoparticle surface in two patterns: physical adsorption or covalent linkage which endow the nanoparticles with diverse biological application.We have adopted the "one-step" method for introducing amine groups onto the nanoparticle surface, namely, by the hydrolysis of silane such as APS and TEOS at the same time. And another route named "two-step" method is also selected for fabricating silica nanoparticles. In this method the addition of APS is executed after the silica nanoparticle was formed. After comparing the fluorescent intensity, morphology, and monodispersity of the products of the two methods, we concluded that the“two-step”method is more preferable for the fabrication of the amine modified silica nanoparticles. Glutaraldehyde, EDC(1-ethyl-3-3(3- dimethylaminopropyl)carbodiimide hydrochloride)/NHS(N-hydroxy-succinimide)are used to couple silica nanoparticles with biomolecules. Glutaraldehyde links the nanoparticles and biomolecules through the Schiff base formed between the amine groups and other biomolecules. By deriving carboxyl groups from the amine groups on the surface, stable amide could be formed between the silica nanoparticles and the recognition molecules with the usage of the coupling agent EDC/NHS. We successfully attached mouse monoclonal antihuman CD71 antibody (McAb CD71) and transferrin onto the nanoparticle surface respectively through the latter method. And cell imaging results affirm that the luminescent nanopartilces successfully labeled the receptors in the fibroblast cells’ membrane. This achievement is helpful for the luminescent silica nanoparticles’ practical application.更多还原