Tedeschi Lorena, Domenici Claudio, Ahluwalia Arti, Baldini Francesco, Mencaglia Andrea
Istituto di Fisiologia Clinica del C.N.R., Pisa e Centro Interdipartimentale di Ricerca E. Piaggio, Università di Pisa, Italy.
Biosens Bioelectron. 2003 Nov 15;19(2):85-93. doi: 10.1016/s0956-5663(03)00173-8.
A study of antibody immobilisation techniques on quartz and fibre optic surfaces for immunosensors has been carried out. Methods of covalent antibody immobilisation which have not previously been applied to optical fibres were investigated, and compared with classical methods found in the literature. Preliminary experiments on covalent immobilisation methods on planar quartz surfaces were conducted to enable us to choose the most suitable protein immobilisation technique for sensor applications. The immobilisation studies were directed in particular towards obtaining a high density of binding sites for the analyte of interest. Two of the most promising methods, antibody immobilisation on surfaces coated with dextran based hydrogel and F(ab')-SH fragments bound to silanised glass, which resulted in surface densities of active sites of above 0.45 pmol/cm2, were selected for further experiments on a fibre optic total internal reflection fluorescence immunosensor and gave satisfactory responses to changes in analyte concentrations of the order of 10(-8) M. The efficiency of polar organic solvents, such as dimethylsulfoxide, in dissociating the antigen-antibody complex and hence to regenerate the immunosensor surface was also evaluated.
开展了一项关于用于免疫传感器的石英和光纤表面抗体固定技术的研究。研究了此前未应用于光纤的共价抗体固定方法，并与文献中发现的经典方法进行了比较。在平面石英表面进行了共价固定方法的初步实验，以便我们为传感器应用选择最合适的蛋白质固定技术。固定化研究尤其旨在获得针对目标分析物的高密度结合位点。选择了两种最有前景的方法，即抗体固定在涂有基于葡聚糖的水凝胶的表面以及与硅烷化玻璃结合的F(ab')-SH片段，这两种方法产生的活性位点表面密度高于0.45 pmol/cm²，并在光纤全内反射荧光免疫传感器上进行了进一步实验，对浓度约为10(-8) M的分析物变化给出了令人满意的响应。还评估了极性有机溶剂（如二甲基亚砜）在解离抗原-抗体复合物从而再生免疫传感器表面方面的效率。
