Abstract

An acetylcholinesterase biosensor has been developed based upon a fourth generation poly(amidoamine) dendrimer with terminal amino groups. The immobilization of acetylcholinesterase was performed by physical adsorption onto the polyelectrolyte complex consisting of dendrimer and poly(styrene sulfonate) or deoxyribonucleic acid as counter ions. The efficiency of immobilization was evaluated by surface plasmon resonance that indicated a twofold increase of the quantity of enzyme on the surface in comparison with similar biosensors with no dendrimer. This property can be attributed to higher roughness of the underlying surface and its possible porosity caused by the star-shaped dendrimer molecules. The influence of the polyelectrolytes used on the biosensor performance as well as kinetic characteristics of enzymatic hydrolysis of thiocholine were assessed. The calibration curves of reversible inhibitors used in the therapy of neurodegenerative diseases (Huperzine A and galantamine) were ascribed with a four-parametrical logistic model. The 15% inhibition levels varied from 9.4 to 65 nM Huperzine A and 90 to 107 nM galantamine depending on the content of the polyelectrolyte complex. The biosensors were tested on example artificial and real urine samples and provided recoveries from 95 to 110%.

摘要

已经基于具有末端氨基的第四代聚（酰胺胺）树状聚合物开发了乙酰胆碱酯酶生物传感器。乙酰胆碱酯酶的固定是通过物理吸附到由树枝状聚合物和聚（苯乙烯磺酸盐）或脱氧核糖核酸作为抗衡离子组成的聚电解质复合物上来进行的。固定效率通过表面等离振子共振进行评估，与没有树枝状聚合物的类似生物传感器相比，表面等离子体共振表明表面上酶的数量增加了两倍。该性质可归因于下表面的较高粗糙度以及由星形树状聚合物分子引起的可能的孔隙率。评估了所使用的聚电解质对生物传感器性能的影响以及硫胆碱酶促水解的动力学特性。用于神经退行性疾病（石杉碱甲和加兰他敏）的可逆抑制剂的校正曲线与四参数逻辑模型相关。根据聚电解质复合物的含量，15％的抑制水平为9.4至65 nM石杉碱甲和90至107 nM加兰他敏。生物传感器在示例性的人工和真实尿液样本上进行了测试，回收率从95％到110％。根据聚电解质复合物的含量，4至65 nM石杉碱A和90至107 nM加兰他敏。生物传感器在示例性的人工和真实尿液样本上进行了测试，回收率从95％到110％。根据聚电解质复合物的含量，4至65 nM石杉碱A和90至107 nM加兰他敏。生物传感器在示例性的人工和真实尿液样本上进行了测试，回收率从95％到110％。