Electrochemiluminescence aptasensor based on bipolar electrode for detection of adenosine in cancer cells

doi:10.1016/j.bios.2013.12.045

Biosensors and Bioelectronics

Volume 55, 15 May 2014, Pages 459-463

https://doi.org/10.1016/j.bios.2013.12.045 Get rights and content

Highlights

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An ECL approach on a wireless ITO bipolar electrode for adenosine detection in cancer cells was provided.
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Ferrocene as DNA tag was introduced on cathodic pole to enhance the ECL of Ru(bpy)₃²⁺/TPA system on anodic pole
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The ECL on the anodic pole could be decreased by the adenosine-induced removal of ferrocene-aptamer on the cathodic pole.
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The approach could detect adenosine in a wide range from 1.0 fM to 0.10 μM.

Abstract

Here we report a novel approach for the detection of adenosine in cancer cells by electrochemiluminescence (ECL) on a wireless indium tin oxide bipolar electrode (BPE). In this approach, ferrocene (Fc) which is labeled on adenosine aptamer is enriched on one pole of the BPE by hybridization with its complementary DNA (ssDNA) and oxidized to Fc⁺ under an external voltage of 5.0 V at the two ends of BPE. Then, a reversed external voltage was added on the BPE, making Fc⁺ enriched pole as cathode. The presence of Fc⁺ promotes the oxidation reaction on the anodic pole of the BPE, resulting in a significant increase of ECL intensity using Ru(bpy)₃²⁺/tripropylamine (TPA) system as test solution. The presence of target adenosine was reflected by the ECL signal decrease on the anodic pole caused by the target-induced removal of ferrocene-aptamer on the cathodic pole. The decrease of ECL signal was logarithmically linear with the concentration of ATP in a wide range from 1.0 fM to 0.10 μM. This ECL biosensing system could accurately detect the level of adenosine released from cancer cells.

Introduction

ECL is an energy-relaxation process by optical emission of an excited molecule produced by an applied potential at an electrode surface of which the excited state of emitting substance is stimulated by electricity instead of chemical reactions. It has become a popular strategy in biological detection for its excellent controllability and lower background noise (He et al., 2013, Li et al., 2007, Li et al., 2011, Tian et al., 2013, Zhang et al., 2013). Recently, ECL detection on BPEs has emerged as a sensitive, high-throughput, and low-cost approach (Chow et al., 2009, Fosdick et al., 2010, Wu et al., 2011, Wu et al., 2013). BPEs are formed by electron conductive wires which are immersed in an ion conductive phase. They are usually set up in microfluidic chips. When a certain voltage is applied between two ends of a microchannel (E_tot), a linear gradient of potential drop will be formed in the electrolyte solution (Mavré et al., 2009, Ulrich et al., 2008a). Since independent conductor behaves like equipotential in conductive solution, a range of interfacial potential differences exist across the surface of the BPE, which is the most significant difference between BPE system and three-electrode configuration (Ulrich et al., 2008b). As a result, the respective interfacial potential difference between the solution and two ends of BPE (E_elec) is a function of axial length of the BPE regardless of the potential drop between external electrodes and the electrolyte solution. The magnitude of ΔE_elec is proportional to the length of the BPE (l_elec) (Chow et al., 2009, Fosdick et al., 2010). Because the rates of the oxidation and reduction reactions are the same at the two ends of the BPE for no electron accumulation is allowed on independent conductor, there is a direct relationship between the rate of the cathodic and anodic reactions. For example, Ru(bpy)₃²⁺ and TPA could be oxidized at the anodic pole of a BPE, and emitting ECL signal, while oxygen would be reduced at the cathodic pole of the BPE at the same time. It was reported that Ru(bpy)₃²⁺ could be oxidized at a lower external voltage when the cathode was dipped in Fe(CN)₆³⁻ (Chang et al., 2012). That is to say, the anodic reactions could be accelerated by the substances which can be reduced easier on the cathodic pole than that of dissolved oxygen.

In this work, we use Fc⁺ to promote the reduction reaction on cathode which correspondingly accelerate the oxidation reactions of Ru(bpy)₃²⁺/TPA system on anode. Based on this, an adenosine biosensor was designed by enriching Fc labeled adenosine aptamer on one pole of the BPE by hybridization with its complementary DNA. In the presence of target adenosine, ECL signal decreased by the target-induced removal of Fc-aptamer on the cathodic pole. Compared with traditional ECL aptasensors, this biosensing platform possesses the advantages of high sensitivity, low-cost, portable sensor system, no need for a direct external connection to the bipolar electrode (wireless detector).

Section snippets

Materials

Ru(bpy)₃²⁺, Bovine serum albumin (BSA), Glutaraldehyde (GA) and TPA were obtained from Sigma-Aldrich. Adenosine triphosphate (ATP), ferrocene modified ATP aptamer (Fc-aptamer) and its complementary DNA (Amino-DNA ) were purchased from Sangon Biotech (Shanghai) Co., LTD. ITO-coated (thickness ~100 nm, resistance ~10 Ω/square) aluminosilicate glass slides were purchased from CSG (Shenzhen, China). Sylgard 184 (including poly (dimethylsiloxane ) (PDMS) monomer and curing agent) was from Dow Corning

Biosensing principle on BPE

In traditional three-electrode system, the reference and counter electrodes functioned only to provide a reference potential and balance the current on the working electrode (Cox et al., 2012). We did most of the time focus on the reaction on working electrode. However a BPE system contained two electrochemical interfaces coupled directly via a conductor. The principles we used in this aptasensor could be divided into two steps: hybridization enhancement (steps a–c) and unlocking helix

Conclusion

In summary, we have successfully fabricated an adenosine aptasensor and realized trace detection of adenosine concentration to the degree of 10⁻¹⁵ M in both standard solutions and actual samples by combing ECL technology with ITO bipolar electrode. Ferrocene on the surface of cathode could accelerate the electron transfer reaction on anode. Faraday current through BPE was increased simultaneously and thus ECL in anode was enhanced. It’s expected that this approach could be used to detect other

Acknowledgments

This work was supported by the 973 Program (Grants 2012CB932600 and 2013CB933800), the National Natural Science Foundation (Grants 21135003, 21305068 and 21105019), and the National Natural Science Funds for Creative Research Groups (Grant 21121091).

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Biosensors and Bioelectronics

Electrochemiluminescence aptasensor based on bipolar electrode for detection of adenosine in cancer cells

Highlights

Abstract

Introduction

Section snippets

Materials

Biosensing principle on BPE

Conclusion

Acknowledgments

Biosensors Bioelectronics

Biosensors Bioelectronics

Electrochem. Commun.

Electrochim. Acta

Biosensors Bioelectronics

Analyst. (London)

J. Am. Chem. Soc.

Anal. Chem.

J. Am. Chem. Soc.

Chem. Commun.

Lab. Chip

Biochemistry

A novel “off-on” ratiometric fluorescent aptasensor for adenosine detection based on FRET between quantum dots and graphene oxide

A bipolar Electrode-Electrochemiluminescence sensor for Salmonella typhimurium detection based on β-cyclodextrin Host-Guest recognition

Paper-based bipolar electrode electrochemiluminescence sensors for point-of-care testing

A novel high throughput electrochemistry corrosion test method: Bipolar electrochemistry

Novel electrochemiluminescent assay for the aptamer-based detection of testosterone

Recent achievements and advances in optical and electrochemical aptasensing detection of ATP based on quantum dots