Challenges of Electrochemical Impedance Spectroscopy in Protein Biosensing
Challenges of Electrochemical Impedance Spectroscopy in Protein Biosensing
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DOI：
10.1021/ac9002358
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发表时间：
2009-05-15
影响因子：
7.4
通讯作者：
Aldissi, M.
中科院分区：
文献类型：
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作者：
Bogomolova, A.;Komarova, E.;Aldissi, M.
在氧化还原剂存在下进行的电化学阻抗谱（EIS）测量是测量在电极表面上发生的电化学非活性化合物的分子相互作用的方便方法。该方法的高灵敏度是非常有利的，也可以与非特异性阻抗变化相关，这种变化很容易被误认为是特异性相互作用。因此，有必要了解所有可能的原因，并进行平行对照实验以排除它们。我们提出的结果在早期阶段的适体为基础的传感器的发展，利用模型系统的人α凝血酶与巯基化的DNA适体相互作用，固定在金电极。EIS测量发生在亚铁氰化物的存在下。除了已知的方法限制，即不能区分特异性和非特异性结合之外，（两者都引起阻抗增加），我们发现了导致非特异性阻抗变化的其他因素，例如：（i）初始电极污染;（ii）重复测量;（iii）额外的循环伏安法（CV）或差分脉冲伏安法（DPV）测量;和（iv）在测量之间在缓冲液中进行另外的孵育，这在以前从未讨论过。我们提出了克服方法局限性的方法。
Electrochemical impedance spectroscopy (EIS) measurement, performed in the presence of a redox agent, is a convenient method to measure molecular interactions of electrochemically inactive compounds taking place on the electrode surface. High sensitivity of the method, being highly advantageous, can be also associated with nonspecific impedance changes that could be easily mistaken for specific interactions. Therefore, it is necessary to be aware of all possible causes and perform parallel control experiments to rule them out. We present the results obtained during the early stages of aptamer-based sensor development, utilizing a model system of human alpha thrombin interacting with a thiolated DNA aptamer, immobilized on gold electrodes. EIS measurements took place in the presence of iron ferrocyanides. In addition to known method limitations, that is, inability to discriminate between specific and nonspecific binding (both causing impedance increase), we have found other factors leading to nonspecific impedance changes, such as: (i) initial electrode contamination; (ii) repetitive measurements; (iii) additional cyclic voltammetry (CV) or differential pulse voltammetry (DPV) measurements; and (iv) additional incubations in the buffer between measurements, which have never been discussed before. We suggest ways to overcome the method limitations.