Optimisation and Characterisation of Anti-Fouling Ternary SAM Layers for Impedance-Based Aptasensors.
Optimisation and Characterisation of Anti-Fouling Ternary SAM Layers for Impedance-Based Aptasensors.
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DOI：
10.3390/s151025015
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发表时间：
2015-09-29
期刊：
影响因子：
--
通讯作者：
Estrela P
中科院分区：
文献类型：
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作者：
Miodek A;Regan EM;Bhalla N;Hopkins NA;Goodchild SA;Estrela P
已经开发了具有增强的防污性能的适体传感器。作为一个案例研究，适体传感器被设计为对人凝血酶具有特异性。该传感平台是在丝网印刷电极上开发的，并且由自组装单层组成，该自组装单层由对人凝血酶特异性的15个碱基的硫醇化DNA适体的三元混合物制成，该适体与1，6-己二硫醇（HDT）共固定并进一步用1-巯基-6-己醇（MCH）钝化。HDT通过其两个硫醇基团形成烷基链桥与表面结合，并且这种结构防止分子非特异性附着到电极表面。使用电化学阻抗谱（EIS），适体传感器能够检测人类凝血酶，因为蛋白质结合后电荷转移电阻（Rct）的变化。在暴露于高浓度的非特异性牛血清白蛋白（BSA）溶液后，未观察到Rct值的变化，突出了所产生的表面的生物污染抗性。在本文中，我们提出了基于三元自组装单层（SAM）层的适体传感器的优化和表征。我们发现，防污性能取决于用于生物传感器建设，这也证实了接触角测量的金表面的类型。我们进一步研究了适体和HDT之间的比例，这可以决定传感层的特异性和选择性。我们还报告了缓冲液的pH值和温度用于孵育的电极与蛋白质的检测和防污性能的影响。最后，通过在不同缓冲液和大气条件下储存修饰电极长达28天来研究适体传感器的稳定性。基于三元SAM层的适体传感器非常有希望用于检测真实的生物样品中的一系列蛋白质的临床应用。
An aptasensor with enhanced anti-fouling properties has been developed. As a case study, the aptasensor was designed with specificity for human thrombin. The sensing platform was developed on screen printed electrodes and is composed of a self-assembled monolayer made from a ternary mixture of 15-base thiolated DNA aptamers specific for human thrombin co-immobilised with 1,6-hexanedithiol (HDT) and further passivated with 1-mercapto-6-hexanol (MCH). HDT binds to the surface by two of its thiol groups forming alkyl chain bridges and this architecture protects from non-specific attachment of molecules to the electrode surface. Using Electrochemical Impedance Spectroscopy (EIS), the aptasensor is able to detect human thrombin as variations in charge transfer resistance (Rct) upon protein binding. After exposure to a high concentration of non-specific Bovine Serum Albumin (BSA) solution, no changes in the Rct value were observed, highlighting the bio-fouling resistance of the surface generated. In this paper, we present the optimisation and characterisation of the aptasensor based on the ternary self-assembled monolayer (SAM) layer. We show that anti-fouling properties depend on the type of gold surface used for biosensor construction, which was also confirmed by contact angle measurements. We further studied the ratio between aptamers and HDT, which can determine the specificity and selectivity of the sensing layer. We also report the influence of buffer pH and temperature used for incubation of electrodes with proteins on detection and anti-fouling properties. Finally, the stability of the aptasensor was studied by storage of modified electrodes for up to 28 days in different buffers and atmospheric conditions. Aptasensors based on ternary SAM layers are highly promising for clinical applications for detection of a range of proteins in real biological samples.