DNA-based biosensors for Hg2+ determination by polythymine-methylene blue modified electrodes

In this work we have developed a new electrochemical DNA-based biosensor for the selective determination of the Hg2+ ion by the use of different electrodes modified with polythymine, bearing methylene blue, as redox probe, in 3' position. The determination of Hg2+ can be employed with an excellent degree of selectivity by the use of DNA biosensors through the formation of the complex Thymine-Hg-Thymine (T-Hg-T): in fact, Hg2+ tends to bind two thymines, generating a T-Hg-T complex with a formation constant higher than that one of the coupling Adenine-Thymine, which can be employed for a selective, fast and cost-effective Hg2+ detection. The presence of the Hg2+ in solution leads to the formation of T-Hg-T complex thus causing the "hairpin-like" folding of oligonucleotide, leading to an improved electronic exchange of methylene blue with the electrode surface due to the reduced distance and thus to an increase of the faradic current which is detected by means of square wave voltammetry (SWV). To test the feasibility of this kind of biosensor to be applied to the analysis of Hg2+ we have developed several biosensors configuration by modifying the electrochemical sensor transducer: (a) Au electrode; (b) Au screen-printed electrode (SPE). The proposed system, allows the determination of Hg2+ in the range 0.2-100nM (0.05-20ppb), with a sensitivity 0.327μA/nM, LOD 0.1nM (0.02ppb), LOQ 0.2nM (0.05ppb) and RSD ≤4.3% when Au electrode is used as electrochemical transducer; on the other hand, in the case of Au SPE the linear range is 0.2-50nM (0.05-10ppb), with a sensitivity 0.285μA/nM, while LOD and LOQ are the same as previously and RSD is ≤3.8%. This enabled the detection of mercury in real samples (waters and fishes) with good accuracy (recoveries 92-101% on waters and 92-107% on fishes, respectively) and reproducibility (RSD ≤9.6% for measurements on waters and ≤8.8% on fishes, respectively).