光纤与生物分子界面上固定FITC标记抗体的荧光光谱

采用化学方法把FITC标记羊抗人抗体IgG共价固定到三氨基三乙氧基硅烷和戊二醛(APTES-Glu)修饰的石英光纤纤芯表面。通过研究未经过任何修饰的纤芯表面的吸附以及共价固定的FITC标记羊抗人抗体的荧光光谱性质发现:共价结合到纤芯表面的FITC荧光光谱相对于溶液中FITC的荧光峰位红移约9nm,而物理吸附的FITC的荧光峰位移动约4nm,而且两者相对荧光强度相差6倍。而在固定的人血清蛋白进行免疫反应后,FITC的荧光峰位只移动3～4nm,除此之外,研究了抗体共价键固定的稳定性问题。结果表明:共价键结合抗体的数量要大于表面吸附,共价键固定的FITC标记羊抗人抗体与光纤表面间的相互作用较吸附于光纤表面的FITC标记羊抗人抗体间的相互作用更强。这对于建立一种通过荧光光谱识别固体表面与生物分子的吸附还是共价结合的判据提供了物理基础。

A Study on Fluorescence Spectroscopy of Immobilizing FITC Labeled Antibody between Fiber Optic Surface and Biomolecule

The interaction of protein with solid surface is not only a fundamental phenomenon but also the key to several applications improtantly and novelly.The immobilization of functional proteins to a solid support is greatly important for biosensors and biochips.In this work,FITC labeled antibody was covalently immobilized on silicon surfaces modified by 3-aminopropyltriethoxysilane(APTES) and glutaraldehyde.The fluorescence spectra of absorption and covalent immobilization FITC labeled antibody was investigated.Subsequently,immobilizing human IgG bound FITC-goat anti-human IgG fluorescence immunoassay were obtained.The results showed that the fluorescence intensity of FITC labeled antibody on covalently immobilized surface was 6 times lager than that on absorption surface.9 nm and 4 nm red-shift at the peak of FITC fluorescence was observed on covalently immobilized surface and adsorption surface,respectively.But 3~4 nm red-shift at the peak of FITC fluorescence was obtained in the fluorescence immunoassay.Finaly,stability of covalently immobilized antibody was studied.The results indicated covalently immobilized antibodies larger than absorbed antibodies on fiber optic surface.And the interaction between immobilizing FITC-labeled antibody and fiber optic surface on covalently immobilization fiber optic surface stronger than that on absorption fiber optic surface.Combining this antibody covalent immobilization methods and the fluorescence spectroscopy analysis technology has the potential to be developed into the fast and simple fluorescence immunoassay and fluorescence multilabelling detection for biosensor.