Electrodes electrochemical detection, nucleic acids

Guanine, with a formal potential at pH 7 of 1.3 V vs. normal hydrogen electrode (NHE) (1.06 V vs. saturated calomel electrode (SCE)), is the most easily oxidized of the four DNA bases/331 The other bases have formal potentials up to 0.5 V more positive. Detecting nucleic acids by electrochemical oxidation depends on structure, with double-stranded (ds) DNA giving only trace oxidation peaks at best and single-stranded (ss) DNA giving irreversible oxidation peaks at about 1.0-1.2 V vs. SCE on carbon electrodes. 

The topics discussed in the book include electrochemical detection of DNA hybridization based on latex/gold nanoparticles and nanotubes nanomaterial-based electrochemical DNA detection electrochemical detection of microorganism-based DNA biosensor gold nanoparticle-based electrochemical DNA biosensors electrochemical detection of the aptamer-target interaction nanoparticle-induced catalysis for DNA biosensing basic terms regarding electrochemical DNA (nucleic acids) biosensors screen-printed electrodes for electrochemical DNA detection application of field-effect transistors to label-free electrical DNA biosensor arrays and electrochemical detection of nucleic acids using branched DNA amplifiers. 

The observation of currents attributable to the faradaic electrochemistry of nucleic acids was pioneered by Palecek and coworkers who studied DNA adsorbed on mercury or carbon electrodes [13]. The signals detected by Palecek were attributable to oxidation of the purines, which produced signals indicative of irreversible processes involving adsorbed bases. These reactions were used as a basis for electrochemical analysis of DNA. Kuhr and coworkers later showed that similar strategies could be developed for analysis of nucleic acids via oxidation of sugars at copper electrodes [14-16]. 

Figure 12.19 Electrochemical detection of thrombin by the interaction between a nucleic acid and a redox-active oligothiophene polyelectrolyte. The electrical contact between the polyelectrolyte and the electrode by means of the aptamer/thrombin complex is blocked. Parts Reprinted with permission from Ref. 66b. Copyright Wiley-VCH Verlag GmbH Co.

The electrochemical behaviour and the adsorption of nucleic acid molecules and DNA constituents have been extensively studied over recent decades [1-6]. Electrochemical studies demonstrated that all DNA bases can be electrochemically oxidized on carbon electrodes [7-13], following a pH-dependent mechanism. The purines, guanine (G) and adenine (A), are oxidized at much lower positive potentials than the pyrimidines, cytosine (C) and thymine (T), the oxidation of which occurs only at very high positive potentials near the potential corresponding to oxygen evolution, and consequently are more difficult to detect. Also, for the same concentrations, the oxidation currents observed for pyrimidine bases are much smaller than those observed for the purine bases. Consequently, the electrochemical detection of oxidative changes occurring in DNA has been based on the detection of purine base oxidation peaks or of the major... 

H. Xie, C. Zhang and Z. Gao, Amperometric detection of nucleic acid at femtomolar levels with a nucleic acid/electrochemical activator bilayer on gold electrode, Anal. Chem., 76 (2004) 1611-1617. 

Fig. 3 Protein detection using immobilized nucleic acids with tethered intercalators. Fabrication of DNA-modified gold electrodes for electrochemical analysis of protein binding and reaction (Reprinted with permission from [174], Copyright(2002) Nature Publishing Group http //www.nature.com/nbt/)...

Fig. 5 Immobilized nucleic acid assays utilizing redox-active moieties, a Amplified detection of viral DNA by generation of a redox-active replica and the bioelectrocatalyzed oxidation of glucose (Reprinted with permission from [200]. Copyright(2002) American Chemical Society), b Alternative formats for the capture on a gold electrode SAM of solution-extended primers or direct surface extension of primer with electrotides (adapted from [185]). c Ferrocene-labelled hairpin for electrochemical DNA hybridization detection. A Fc-hairpin-SH macromolecule is immobilized on a gold electrode. When a complementary DNA target strand binds to the hairpin, it opens and the ferrocene redox probe is separated from the electrode, producing a decrease in the observed current (Reprinted with permission from [203], Copyright(2004) American Chemical Society)...

An electrochemical DNA hybridization biosensor basically consists of an electrode modified with a single stranded DNA called probe [109]. Usually the probes are short oligonucleotides (or analogues such as peptide nucleic acids). The first and most critical step in the preparation of an electrochemical DNA biosensor is the immobilization of the probe sequence on the electrode. The second step is the hybrid formation under selected conditions of pH, ionic strength and temperature. The next step involves the detection of the double helix... 

Enzyme DNA hybridization assays with electrochemical detection can offer enhanced sensitivity and reduced instrumentation costs in comparison with their optical counterparts. Efforts to prevent non-specific binding of the codissolved enzyme and to avoid fouling problems by selecting conditions suitable to amplify the electrode response have been reported by Heller and co-workers [107]. A disposable electrochemical sensor based on an ion-exchange film-coated screen-printed electrode was described by Limoges and co-workers for an enzyme nucleic acid hybridization assay using alkaline phosphatase [108] or horseradish peroxidase [109]. In another methodology to improve sensitivity, a carbon paste electrode with an immobilized nucleotide on the electrode surface and methylene blue as hybridization indicator was coupled, by Mascini and co-workers [110], with PGR amplification of DNA extracted from human blood for the electrochemical detection of virus. 

Figure 5-3. Electrocatalytic nucleic acids detection based on the reduction of FefCNjg mediated by RufNHj) . The Ru(III) complex is electrostatically bound to the anionic backbone of an immobilized DNA oligonucleotide and upon the hybridization of a complementary target sequence is present at a higher concentration at the electrode surface. This increases the surface concentration of the redox mediator and leads to an enhanced electrochemical signal tliat can be used to detect specific DNA sequences. The presence of Fe(CN)6 chemically regenerates electrochemically reduced Rufll), and thereby catalytically amplifies the signals. (Adapted from ref. 11).

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