Electroactive DNA

The reported strategies utilized in DNA sensing include (1) sequence-specific hybridization processes based on the oxidation signal of most electroactive DNA bases, guanine and adenine [13,24] or (2) quasi-specific detection of small molecules capable of binding by intercalation or complexation with DNA, such as metal coordination complexes, antibiotics, pesticides, pollutants, etc. [17,18] or in the presence of some metal tags such as gold, silver nanoparticles, etc. [23,50,51]. 

Numerous electrochemical strategies have been developed for the detection of mismatches in DNA. These vary from the use of electroactive DNA intercalators to enzymatic signal amplihcation schemes, or redox-modified oligonucleotides. In the following sections, we will focus on the discussion of a range of electrochemical mismatch detection schemes. 

Label-free A decrease/increase in the oxidation/reduction peak current of electroactive DNA bases such as guanine or adenine is monitored. 

Electrochemical biosensors have received considerable interest in specific DNA detection. The first reports about the electrochemistry of DNA were published by the end of the 1950s and in the beginning of the 1960s by Palecek. These reports were based on electrochemical reduction and oxidation signals of DNA due to the electroactive DNA bases. 

Electrochemical detection of successful DNA hybridization events should be also considered. Although it is based mostly on external electrochemical markers, such as electroactive indicators or enzymes, the exploitation of the intrinsic DNA oxidation signal requires a multi-site attachment such as adsorption as the immobihzation technique. 

This conclusion is also supported by the fact that, in contrast to ssDNA, the oxidation signal coming from dsDNA is poorly developed at both GC and GC(ox). This is probably attributable to the electroactive A and G residues in dsDNA being inaccessible to the surface, while most bases in denatured DNA can freely interact with the GC(ox) surface. On the other hand, the hydrogen-bonded bases in native DNA are hidden within the double heUx, a serious steric barrier to electron transfer between the purine and the GC(ox). 

Differential pulse voltammetry and electrochemical impedance have demonstrated that G, A, guanosine, and their oxidation products are electrostatically adsorbed on GC and GC(ox) surfaces [47,49]. The strength of adsorption of the DNA bases on the GC surface were found to be similar [49]. Strongly adsorbed G dimers were formed on GC between G and the adsorbed G oxidation products, which slowly cover and block the surface. The appHcation of ultrasound led to removal of the adsorbed species. The effect of this was mainly to enhance transport of electroactive species and to clean the electrode in situ, avoiding electrode fouling. 

Electropolymerizable monomers that give rise to conducting electroactive polymers (CEPs) or insulating polymer thin films provide a convenient approach for the immobilization of DNA. More importantly, this method provides an easy means to achieve spatial separation of the ssDNA sequence... 

Another coupling method, i.e. cross-linking or entrapment in polymeric films, which has been used to create a more permanent nucleic acid surface, is described in some chapters (e.g. conductive electroactive polymers for DNA immobilization and self-assembly DNA-conjugated polymers). One chapter reviews the basic characteristics of the biotin-(strept)avidin system laying the emphasis on nucleic acids apphcations. The biotin-(strept)avidin system can be also used for rapid prototyping to test a large number of protocols and... 

A disposable electrochemical enzyme-amplified genosensor was described for specific detection of Salmonella (Del Giallo et al., 2005). A DNA probe specific for Salmonella was immobilized onto screen-printed carbon electrodes and allowed to hybridize with a biotinylated PCR-amplified product of Salmonella. The hybridization reaction was detected using streptavidin conjugated-AP where the enzyme catalyzed the conversion of electroinactive a-naphthyl phosphate to electroactive a-naphthol, which was detected by differential pulse voltammetry. 

K.M. Millan and S.K. Mikkelsen, Sequence-selective biosensor for DNA based on electroactive hybridization indicators, Anal. Chem., 65 (1993) 2317-2323. 

The formation of double-stranded DNA upon hybridisation is commonly detected in connection with the use of an appropriate electroactive hybridisation intercalator or labelling DNA by a simple electroactive molecule or an adequate NP. 

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