Graphene materials have also been employed for sensitive and selective electrochemical detection of nucleobases, nucleotides, single stranded DNAs (ssDNA), and double stranded DNAs (dsDNA). Such electrochemical DNA sensors may provide a simple alternative approach for DNA analysis and sequencing.
The four distinct nucleobases (A: adenine, T: thymine, C:cytosine, G: guanine) can be electrochemically differentiated because they have different oxidation potentials. Demonstrations using RGO with abundant –COOH groups to electrochemically detect guanine and adenine with a LOD of 50 nM and 25 nM, respectively has been reported in the literature. The high sensitivity can be ascribed to the excellent electrochemical properties of RGO, the electrostatic attraction between the negatively charged –COOH groups and the positively charged nucleobases, and the strong p–p stacking interaction between the nucleobases and honeycomb carbon lattice.
A Fe3O4 nanoparticle doped RGO–chitosan electrode has been used to detect guanosine. It was suggested that Fe3O4 nanoparticles help to reduce the electron transfer resistance.
RGO based electrode decorated with AuNPs by potentiostatic electro deposition to detect ssDNA has been shown in the literature. The incorporation of AuNPs was proven to be essential to separate the oxidation signal of T from that of A. It was also demonstrated that the electrochemically reduced RGO showed enhanced electrochemical and electro catalytic activity as compared to chemically reduced RGO. This DNA sensor is able to detect single-base alteration (mutation) without any labeling or probe DNA.
Stacked graphene nanofibers (SGNFs) were used to distinguish the four nucleobases with a sensitivity two to four folds higher than carbon nanotube-based electrodes. The high sensitivity is due to numerous open edges of individual graphene nanosheets which are much more electrochemically active compared to the basal carbon plane. This sensor was employed to examine the base composition of human influenza A(H1N1) DNA strand.
Graphene epitaxially grown on SiC was used to detect dsDNA was also demonstrated in the literature. It was shown that dsDNA can be differentiated from ssDNA, because dsDNA exhibits lower oxidation peaks for A and C and increased oxidation potential for C. Electrochemical detection of dsDNA is not possible with the conventional electrodes (e.g., gold electrode and GCE) due to their limited electrochemical potential window. It was also shown that electrochemical anodization to introduce oxygenated groups onto graphene largely improved the electrode performance.
A GO modified electrode was used for detection of DNA hybridization. In this work, probe ssDNA molecules that lack guanine base were covalently immobilized onto a GO film, and hybridization was detected by the guanine oxidation signal from the target ssDNA molecules (a hepatitis B virus specific sequence).
An interesting demonstration showed RGO quantum dots (B10 nm) used to modify the pyrolytic graphite electrode for detection of DNA hybridization. When the target ssDNA hybridizes with the pre-immobilized probe ssDNA, the electron transfer from the electrochemically active species [Fe(CN)6]3-/4- was increased because the blocking effect by the probe ssDNA was alleviated. A LOD of 100 nM was reached. This study suggests the potentials of RGO quantum dots in electrochemical sensing. The good performance of RGO quantum dots may be attributed to their abundant edge sites (electrochemically active sites) and quantum confinement effects.
Based on a similar sensing scheme, a RGO based sensor to detect hybridization of methicillin-resistant Staphylococcus aureus DNA with a LOD of 100 fM was shown. However, the scientists proposed an opposite mechanism. They argued that hybridized DNAs remained on the RGO surface and caused an increase of electron transfer resistance (hence a decrease in the electrochemical signal). The discrepancy between both works may be because of the size difference between RGO quantum dots and RGO sheets. Larger RGO sheets likely can bind more strongly with hybridized DNAs.
Hypoxanthine is a purine derivative. A hypoxanthine sensor was constructed using an electrode consisting of RGO, conducting polypyrrole graft copolymer, poly(styrenesulfonic acid-g-pyrrole), and enzyme xanthine oxidase. The detection mechanism of such a sensor involved two-steps of oxidation: oxidation of hypoxanthine catalysed by xanthine oxidase, and subsequent oxidation of uric acid and H2O2 produced from the previous reaction. RGO and the conducting polymer interact with p–p stacking and form a nanocomposite with high conductivity and an excellent electrocatalytic environment. As a result, a LOD of 10 nM was obtained. As hypoxanthine accumulates continuously from adenine nucleotide degradation after fish death, this sensor was employed to assess fish freshness.
Figure: Graphene-based electrochemical DNA sensor. (a) Schematics of graphene sheet orientation in multiwalled carbon nanotubes (upper) and stacked graphene nanofibers (lower). The highly electroactive edge portion of the sheets is represented in yellow. (b) Differential pulse voltammetry (DPV) for ssDNA of the human influenza A(H1N1) obtained from SGNF (stacked graphene nanofibers, red), GMP (graphite microparticle, green), MWCNT (multi-walled carbon nanotubes, blue), GC (glassy carbon, black dashed), and EPPG (edge plane pyrolytic graphite, black dotted) electrodes.
Extracted and edited from “Biological and chemical sensors based on graphene materials by Yuxin Liu, Xiaochen Dong and Peng Chen in Chemical Society Reviews, 2012″