Preparation and characterization of zinc oxide nanoparticles and their sensor applications for electrochemical monitoring of nucleic acid hybridization

https://doi.org/10.1016/j.colsurfb.2011.04.030Get rights and content

Abstract

In this study, ZnO nanoparticles (ZNP) of approximately 30 nm in size were synthesized by the hydrothermal method and characterized by X-ray diffraction (XRD), Braun–Emmet–Teller (BET) N2 adsorption analysis and transmission electron microscopy (TEM). ZnO nanoparticles enriched with poly(vinylferrocenium) (PVF+) modified single-use graphite electrodes were then developed for the electrochemical monitoring of nucleic acid hybridization related to the Hepatitis B Virus (HBV). Firstly, the surfaces of polymer modified and polymer–ZnO nanoparticle modified single-use pencil graphite electrodes (PGEs) were characterized using scanning electron microscopy (SEM). The electrochemical behavior of these electrodes was also investigated using differential pulse voltammetry (DPV) and electrochemical impedance spectroscopy (EIS). Subsequently, the polymer–ZnO nanoparticle modified PGEs were evaluated for the electrochemical detection of DNA based on the changes at the guanine oxidation signals. Various modifications in DNA oligonucleotides and probe concentrations were examined in order to optimize the electrochemical signals that were generated by means of nucleic acid hybridization. After the optimization studies, the sequence-selective DNA hybridization was investigated in the case of a complementary amino linked probe (target), or noncomplementary (NC) sequences, or target and mismatch (MM) mixture in the ratio of (1:1).

Highlights

ZnO nanoparticles around 30 nm diameter were hydrothermally synthesized. ► The particles were characterized by X-ray diffraction, Braun-Emmet-Teller N2 adsorption analysis and transmission electron microscopy. ► ZNPs enriched poly(vinylferrocenium) modified graphite electrodes were developed as the first time in our study for selective and sensitive recognition of nucleic acids.

Introduction

Electrochemical sensors are playing a growing role in various fields where accurate, low-cost, rapid, and on-line measuring systems are required. In particular, these devices offer elegant routes for interfacing, at the molecular level, biological recognition events and electronic signal transduction processes. In addition to these properties, electrochemical biosensors offer several advantages such as simpler immobilization of the DNA layer (i.e. adsorption driven by a positive electrode potential) and faster measurement times. Rapid testing of nucleotide sequences is required in different fields, including disease diagnostics, genetic testing, forensic medicine, rapid detection of biological warfare agents, environmental testing, drug–DNA interaction, etc. [1], [2], [3], [4], [5]. The development of inexpensive, easy-to-use, and rapid measurement analytical devices, therefore, has been the focus of numerous research efforts. Combining the advantages of electrochemical biosensors with nanotechnology plays a fundamental role in the design and development of electrochemical nanobiosensors due to their high selectivity and sensitivity [6], [7]. Indeed, there is great interest in electrochemical studies of semiconductor materials that offer the distinct advantages of biomolecular compatibility, chemical stability as well as the straightforward synthesis of nanomaterials with controlled dimensionality, morphology, biocompatibility, and enhanced electrochemically active surfaces [8], [9].

Electrochemical DNA hybridization biosensors exploit the use of different nanomaterials such magnetic particles, nanoparticles, and carbon nanotubes [10], [11], [12], [13]. Electrochemistry provides an attractive route to achieve specific DNA detection in a simple, rapid and inexpensive manner [14], [15], [16], [17], [18], [19].

Zinc oxide (ZnO) nanoparticles represent a recent topic of research due to their wide band gap and large excitation energy. Various nanostructures can be formed with this material: nanowires, nanotubes, nanorods, nanoribbons, nanoneedles, nanocables, etc. [20]. There is increased demand for the usage of ZnO nanoparticles in many devices such as solar cells, batteries, photodetectors, nanolasers and biosensors [21], [22], [23], [24], [25], [26], [27], [28], [29], [30], [31]. The electrochemical behavior of hemoglobin entrapped in Nafion/nano-ZnO film on the surface of an ionic liquid-modified carbon paste electrode was investigated by Sun et al. [26]. Liu et al. fabricated carbon-decorated ZnO nanowire arrays for direct electrochemistry of enzymes (glucose oxidase, horseradish peroxidase) and biosensing applications [27]. Flower-like ZnO–gold nanoparticles (GNPs)–Nafion nanocomposites were used to promote direct electron transfer with horseradish peroxidase (HRP) [28]. Amperometric glucose biosensors formed the basis of aligned ZnO nanorod films studied by Liu et al. [29]. Nanoporous ZnO films were prepared on graphite electrodes for myoglobin (Mb) immobilization by Zhao et al. [30]. A biosensor based on ZnO nanorod clusters was utilized as a platform for the immobilation of tyrosinase on nanocrystalline diamond electrodes [31].

In this study, ZnO nanoparticles were successfully synthesized by the hydrothermal method. The particles were characterized by X-ray diffraction (XRD), Braun–Emmet–Teller (BET) N2 adsorption analysis and transmission electron microscopy (TEM). ZnO nanoparticles (ZNP)–poly(vinylferrocenium) (PVF+) modified single-use pencil graphite electrode (PGE) was then developed for use as an electrochemical DNA hybridization sensor. Various modifications of the PGE were characterized by scanning electron microscopy (SEM) and electrochemical impedance spectroscopy (EIS). The electrochemical behavior of these electrodes was also investigated using differential pulse voltammetry (DPV). ZNP–PVF+ modified PGEs were tested for the electrochemical detection of DNA based on the changes at the guanine oxidation signals. Various modifications in DNA oligonucleotides and probe concentrations were examined in order to optimize the electrochemical signals that were generated by means of nucleic acid hybridization. After the optimization studies, the sequence-selective DNA hybridization was investigated in the case of complementary of amino linked probe (target), or noncomplementary (NC) sequences, or target and mismatch (MM) mixture in the ratio of (1:1).

Section snippets

Apparatus

The X-ray diffraction (XRD) analysis of the nanoparticles was executed using a Rigaku D/Max – 2200 ULTIMAN X-ray diffractometer using Cu Kα radiation (λ = 1.5418 Å). The transmission electron micrographs (TEM) were taken using an FEI Tecnai G2 transmission electron microscope (at 200 kV). The BET surface area of the nanoparticles was determined using a Quantachrome Corporation Autosorb-1-C/MS.

All voltammetric measurements were performed with an AUTOLAB-PGSTAT 302 electrochemical analysis system

Results and discussion

Fig. 1 shows the XRD pattern of ZnO nanoparticles. The figure reveals peaks that mainly correspond to zincite with a hexagonal structure. The Scherrer equation was used, assuming a shape factor of 0.9, to estimate the average crystalline size in conjunction with the ZnO reflections:D=0.9λβcosθwhere λ is 0.15418 nm for Cu-Kα, β is the Bragg diffraction angle and θ is the full width at half maximum of the diffraction peak (FWHM).

Furthermore, it is known that FWHM can be interpreted in terms of

Conclusions

In this study, nanoscale ZnO particles with around 30 nm diameter were hydrothermally synthesized. The particle size, confirmed by TEM analysis, was with the value calculated from the Scherrer equation. XRD analysis of the particles in conjunction with the database of both instruments confirms that the synthesized nano-sized ZnO powder had a purely tetragonal zincite structure. We reported, for the first time, the electrochemical monitoring of nucleic acid hybridization relating to the detection

Acknowledgements

A.E. expresses her gratitude to the Turkish Academy of Sciences (TUBA) as an Associate member for their support.

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