Electrochemical Aptamer-Based Biosensors

Dipartimento di Chimica, University degii Studi di Firenze, Via della Lastruccia 3, 50019 Sesto Fiorentino, and Istituto Nazionaie Biostrutture e Biosistemi (INBB), 

Viale Medagiie d Oro 305, 00136 Roma, Itaiy marco.mascini unifi.it 

Aptamers are oligonucleotides (DNA or RNA molecules) which are able to bind selectively to low-molecular-weight molecules or to macromolecules such as proteins. The interest in aptamers as ligands is related to their ease of preparation by an evolutionary selection procedure that eliminates the need for structural design of the ligand sites. Selection of the aptamers for the specific target is based on the SELEX (systematic evolution of ligands by exponential enrichment) procedure. 

In recent years, great progress has been made toward the development of aptamer-based assa. These assays can be set up in a wide variety of formats (direct, sandwich, or competitive assays), which are summarized in Fig. 2.1. 

The potential use of aptamers as receptors in biosensors and bioassays has been extensively reviewed [1-7] and also several books have appeared in the last years [8,9]. In this chapter, the current status of research in electrochemical aptasensors is considered. Attention is focused on label-free and labeled aptasensors, and the anal3Aical capabilities of these devices are discussed. 

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Han K, Chen L, Lin ZS et al (2009) Target induced dissociation (TID) strategy for the development of electrochemical aptamer-based biosensor. Electrochem Commun 11 157-160... 

Radi, A.-E., 2011. Electrochemical aptamer-based biosensors recent advances and perspectives. Int. J. Electrochem. 2011, 1-17. 

A very recent example of the use of this redox probe in an aptamer-based biosensor was published by Kim et al. [33]. An electrochemical biosensor for ox3d etracycline detection was developed using ssDNA aptamer immobilized on gold interdigitated array (IDA) electrode chip (Fig. 2.7). Cyclic voltammetry and square wave voltammetry were used to measure the current at the electrode chip... 

E. E. Ferapontova and K. V. Gothelf, Optimization of the electrochemical RNA-aptamer based biosensor fo theophylline by using a methylene blue redox label. Electroanalysis, 21, 261-1266 [2009a],... 

Aptamer recognition has been described for the development of biosensors and applied to biomedical and environmental studies [46]. As illustrated in Fig. 5.7, an electrochemical aptamer-based sensor (E-AB) for specific recognition of thrombin was constructed. Zhang and co-workers [46] used immobilization of a Fe304-nanoparticles/tagged aptamer via a self-assembly method. In this case, bifunctional aptamer was covalently linked to both Fe304-NPs and gold electrode. Certainly... 

Barthelmebs, L., Hayat, A., Limiadi, A.W. et al. (2011) Electrochemical DNA aptamer-based biosensor for OTA detection, using superparamagnetic nanoparticles. Sens. Actuators, B, 156, 932-937. 

Aptamer-based biosensors, also called aptasensor have gain a wide interest in the last years due to the advantages of aptamers compared to antibodies. Similar to antibodies, a variety of immobilization methods is available to bind aptamers to the sensor element. Aptasensors can be coupled to an electrochemical, optical or mass-sensitive transducer [13]. One of the successful examples for aptasensor was the detection of thrombin which was widely investigated [14]. Xiao et al. [15] have made an interesting development a redox compound (methylene blue) was inserted into the thrombin aptamer. When the target bound to the aptamer, the induced conformation change inhibited the electron transfer from the methylene blue to the electrode. This change could be detected amperometrically. 

In this part we will describe recent achievements in the development of biosensors based on DNA/RNA aptamers. These biosensors are usually prepared by immobilization of aptamer onto a solid support by various methods using chemisorption (aptamer is modified by thiol group) or by avidin-biotin technology (aptamer is modified by biotin) or by covalent attachment of amino group-labeled aptamer to a surface of self-assembly monolayer of 11-mercaptoundecanoic acid (11-MUA). Apart from the method of aptamer immobilization, the biosensors differ in the signal generation. To date, most extensively studied were the biosensors based on optical methods (fluorescence, SPR) and acoustic sensors based mostly on thickness shear mode (TSM) method. However, recently several investigators reported electrochemical sensors based on enzyme-labeled aptamers, electrochemical indicators and impedance spectroscopy methods of detection. 

Label-free protein biosensors witnessed great advance in recent years. An impedometric label-free detection was proposed to conduct immunoassay with high sensitivity and specificity [140]. Aptamers that were immobilized on solid substrates received appreciable attention because they have high permanent charge density, which is possibly exploited for label-free detection such electrochemical impedance spectroscopy (EIS) [141]. There are many cases of aptamers-based sensors, which are separately described in Sect. 4.4. 

Hansen, J. A., Wang, J., Kawde, A-N., Xiang, Y., Gothelf, K. V., CoUins, G. (2006). Quantum dot/aptamer-based ultrasensitive multi-analyte electrochemical biosensor. J Am Chem Soc 128, 2228-2229. 

In its simplest, QCM, format, protein-aptamer interactions were analyzed by Liss et al. (2002). They compared the interaction of IgE with DNA aptamer as well as with anti-IgE antibodies. Although the detection limit was similar in the two cases, the advantage of the aptasensor was its possibility of surface regeneration, which was impossible for an antigen-based biosensor. However, recently it has been shown that immobilization of anti-IgE on the dendrimer surface also allows us to regenerate an immunosensor (Svobodova et al., 2006). The QCM method was recently compared with the electrochemical biosensor assay of thrombin detection (Hianik et al., 2005, 2007). It has been shown that the sensitivity of thrombin detection was similar for the two methods. Mascini and co-workers showed that similar results in sensitivity and selectivity in the detection of Tat peptide with RNA aptamer can be obtained by the QCM and SPR methods (Tombelli et al., 2005b). 

In this chapter several applications of aptamers in the development of electrochemical biosensors have been reported. Different electrochemical methods based on aptamers have been considered both for the detection of proteins (PDGF, VEGF, lysozyme, or thrombin) or small molecules. 

F. Wei and C. M. Ho, Aptamer-based electrochemical biosensor for Botulinum neurotoxin, Ano/. Bioanai Chem., 393,1943-1948 [2009]. 

This chapter covers recent advances in the development of DNA electrochemical biosensors making use of gold nanoparticles to improve their anal5d ical performance. Aptamer-nanoparticle-based biosensors will be also covered since using exponential selection strategies, various RNA and DNA sequences have been identihed that bind small molecules and proteins while inducing a change... 

Deng CY, Chen JH, Nie LH et al (2009) Sensitive bifimctional aptamer-based electrochemical biosensor for small molecules and protein. Anal Chem 81 9972-9978... 

Ferapontova EE, Olsen EM, Gothelf KV. An RNA aptamer-based electrochemical biosensor for detection of theophylline in serum. Journal of the American Chemical Society 2008 130 4256-8. http //dx.doi.org/10.1021/ja711326b. 

He, P., Oncescu, V., Lee, S., Choi, L, Erickson, D., 2013. Label-free electrochemical monitoring of vasopressin in aptamer-based microfluidic biosensors. Analytica Chimica Acta 759, 74-80. 

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