2300 Current Organic Chemistry, 2010, 14, 2300-2309 1385-2728/10 $55.00+.00 © 2010 Bentham Science Publishers Ltd. Recent Advances of Sensitive Electroanalytical Tools and Probes in the Study of DNA Structure S. Girousi*, C. Serpi, S. Karastogianni and A. Ioannou Analytical Chemistry Laboratory, School of Chemistry, 541 24 Thessaloniki, Greece Abstract: The research and development of electrochemical biosensors applied to life sciences during the past years were reviewed in the present article. In the first part of the article, a brief introduction on nanomaterial based electrochemical DNA biosensors is given. In the second part DNA biosensors and transition metal complexes with biological interest are described. Finally, the development trends of electrochemical biosensors in genetic and epigenetic testing are being discussed. Keywords: Electrochemical biosensors, electroanalysis, nanomaterials, DNA interactions, methylated DNA. NANOMATERIAL – BASED ELECTROCHEMICAL DNA BIOSENSORS In recent years, different nanostructured materials have begun to be used widely in the preparation of DNA biosensors. Nanomate- rials are characterized by at least one dimension smaller than 100 nm and they display unique physical and chemical features due to the quantum size effect, mini size effect, surface effect and macro- quantum tunnel effect [1]. The use of nanomaterials, due to their excellent mechanical, optical, electrical and thermal properties could significantly improve the sensitivity of electrochemical bio- sensors [2]. The electrochemical nanobionsensors were applied in areas of cancer diagnostics and detection of infectious organisms [3]. Materials like metal (gold, silver), carbon and polymers (espe- cially conducting polymers) have been used to prepare nanomateri- als such as nanoparticles, nanotubes and nanowires [4]. Carbon nanotubes (CNTs) are one of the most important group of nanomaterials. CNTs, can be considered as rolled up graphite sheets held together by Van der Waal’s bonds [5]. CNTs include both singlewalled (SWNT) and multiwalled (MWNT) structures [6]. SWNTs consist of a single CNT with a typical diameter in the range of 0.4 to 2nm and a length up to few micrometers. The MWNTs which are formed from several concentric CNTs, which have diameters that normally exceed 2nm, while the lengths may be more than 10 mm [7]. CNTs have attracted much attention as analytical tools due to their special chemical, electrical and mechanical properties. Chemi- cal reactivity of nanotubes, is compared with that of graphene sheet, enhanced as a direct result of the curvature of the CNT surface [2]. Depending on their chirality, CNTs can act as metals or as semi- conductors. Usually, MWNTs are regarded as metallic conductors. In contrast, SWTNs may possess different chirality and they can be either metals or semiconductors. Chiral nanotubes have metallic properties, whereas arm-chair and zig-zag nanotubes have the prop- erties of semi-conductors [7]. For the analytical applications the lack of solubility of CNTs, due to the hydrophobicity, is a major problem [7]. Strategies for CNTs dispersion / solubilization can be classified into in three types, namely [8]: (i) dispersion upon oxidative acid *Address correspondence to this author at the Analytical Chemistry Laboratory, School of Chemistry, 541 24 Thessaloniki, Greece; Tel: ---------------; Fax: ----------------------; E-mail: girousi@chem.auth.gr treatments (HNO 3 , HNO 3 / H 2 SO 4 mixture), (ii) non-covalent stabi- lization (sodium dodecylbenzene sulfonate (SDBS), sodium dode- cyl sulfate (SDS), triton X-100 ,-cuclodextrin ) and; (iii) covalent stabilization ( glucose, DNA, enzymes) . A very critical step for using CNTs in biosensor development is their treatment [9]. Many methods have been reported for the CNTs modification, including [10, 11]: (a) treatment with acids or bases for activating the CNTs and/or increasing the micropore volume of the CNTs. In that case, the surface of CNTs can be generated with high concentrations of carboxylic, carbonyl and hydrohyl groups. (b) Modification of the CNTs with conducting polymers such as polypyrrole or transition-metal oxides. For CNT-based DNA biosensors, DNA is generally immobi- lized at CNTs by two approaches: non covalent attachment (psychi- cal adsorption and entrapment) and covalent binding [12]. Carbon paste and other composite electrodes present a large number of advantages over other working electrodes, like low background currents and easy renewal [7]. For that reason compos- ites based on the use of carbon nanotubes have received much at- tention [13]. A variety of binders such as mineral oil or nujol, Tef- lon and bromoform can be used to produce CNT pastes or compos- ites [7]. Pedano and co-workers [14] studied the adsorption and the electrooxidation of free guanine and adenine, oligonucleotides and polynucleotides at carbon nanotube paste electrodes (CNTPE) using adsorptive stripping potentiometric techniques. The CNTPE were prepared by mixing multi-walled carbon nanotubes powder and mineral oil. They concluded that (1) free guanine can be adsorbed at CNTPE under conditions that at a classical (grarhite) carbon paste electrode (CPE) could not be adsorbed. Adenine, could be adsorbed at both carbon electrodes, especially at CNTPE and more strongly at the pretreated one (2). Trace (μg/L) levels of the oli- gonucleotides and polynucleotides could be readily detected follow- ing short accumulation periods with detection limits of 2.0 μg / L for a 21 bases oligonucleotide and 170 μg / L for calf thymus dsDNA (3). The interaction between nucleic acids and CNTPE presented mainly a hydrophobic character and the confined nucleic acid layers demonstrated to be stable in air. A paste electrode assembled by MWCNTs has been developed by Qi et al. [15]. DNA probe was immobilized on the electrode’s surface with polypyrrole electropolymerization with high sensitivity and selectivity for the detection of DNA hybridization. The detec- tion approach is supported by the changes in the current signal gen-