Laboratory of Analytical Chemistry, Department of Chemistry, University of Crete Voutes, P.O. Box 2208, 71003 Iraklion, Crete, Greece.
Anal Bioanal Chem. 2010 Dec;398(7-8):3015-21. doi: 10.1007/s00216-010-4253-z. Epub 2010 Oct 9.
CdSe/ZnS core/shell quantum dots (QDs) are functionalized with mercaptoundecanoic acid (MUA) and subsequently covered with poly-L-lysine (PLL) as the template for the formation of the silica outer shell. This nanocomposite is used as a transduction and stabilization system for optical biosensor development. The covalent immobilization of the enzyme acetylcholinesterase from Drosophila melanogaster (AChE) during the formation of the biomimetically synthesized silica is used here as a model, relatively unstable enzyme, as a proof of principle. The enzyme is successfully immobilized onto the QDs and then stabilized by the PLL capping and the subsequent formation of the outer nanoporous silica thin shell, giving rise to the QD/AChE/PLL/silica biosensor. It is shown that the poly-L-lysine templated silica outer shell does not modify the optical properties of the quantum dots, while it protects the enzyme from unfolding and denaturation. The small pores of the silica shell allow for the free diffusion of the analyte to the active center of the enzyme, while it does not allow for the proteases to reach the enzyme. The response of the QD/AChE/PLL/silica nano-biosensor to its substrate, acetylcholine chloride, is evaluated by monitoring the changes in the QDs' photoluminescence which are related to the changes in pH. These pH changes of the surrounding environment of the QDs are induced by the enzymatic reaction, and are associated with the analyte concentration in the solution. The biodetection system proposed is shown to be stable with a storage lifetime of more than 2 months. The data presented provides the grounds for the application of this nanostructured biosensor for the detection of AChE inhibitors.
硒化镉/硫化锌核/壳量子点（QDs）用巯基十一酸（MUA）功能化，然后用聚-L-赖氨酸（PLL）覆盖作为形成二氧化硅外壳的模板。这种纳米复合材料被用作光学生物传感器开发的转导和稳定系统。在这里，使用相对不稳定的酶——黑腹果蝇乙酰胆碱酯酶（AChE）的共价固定化作为模型，在仿生合成的二氧化硅形成过程中，作为一个原理证明。酶成功地固定在量子点上，然后通过 PLL 封端和随后形成的外部纳米多孔二氧化硅薄壳进行稳定，从而产生 QD/AChE/PLL/二氧化硅生物传感器。结果表明，多聚-L-赖氨酸模板化的二氧化硅外壳不会改变量子点的光学性质，同时保护酶免于展开和变性。二氧化硅壳的小孔允许分析物自由扩散到酶的活性中心，同时不允许蛋白酶到达酶。QD/AChE/PLL/二氧化硅纳米生物传感器对其底物——氯化乙酰胆碱的响应通过监测与 pH 值变化相关的量子点光致发光的变化来评估。这些 QD 周围环境的 pH 值变化是由酶反应引起的，与溶液中分析物的浓度有关。所提出的生物检测系统显示出稳定性，存储寿命超过 2 个月。所提供的数据为该纳米结构生物传感器在检测 AChE 抑制剂方面的应用提供了依据。
