Electrochemical Biosensors Based On In-situ Growth Of Metal Nanometerials On The DNA Modified Interfaces

Electrochemical biosensor is a new subject and technology, which comes from the combination and growth of many subjects, such as chemistry, biology, physics, electronic technology and so on. Because of its simple to operate, high sensitivity and real-time measuring, electrochemical biosensor has important applications in some areas such as clinic medicine, environment measurement and food industry, also has been focused in research. Nanoparticles have small dimension effect, quanta dimension effect, surface effect and macroscopical quanta tunnel effect, so they manifest a series of specific physical and chemical characters, and provides new thinkings and routes for the construction of electrochemical DNA biosensor. In this paper, metals nanoparticles were in-situ growth on the DNA modified interfaces to composited electrochemical sensors, and the concrete research content are as follows:(1) A high sensitivity DNA electrochemical biosensor was constructed based on in-situ growth of double-stranded DNA templated copper nanoclusters(ds DNA-templated Cu NCs) on the DNA modified interfaces. The Au electrode(Au E) were modified with DNA probes first, and then treated with 6-mercaptohexanol(MCH) to block the non-specific sites on the surface. When the probes DNA hybridized with the target DNA to form a double stranded structure via the specific interaction, the electroactive ds DNA-templated Cu NCs formed specifically by the disproportionation of Cu Br at the big groove of ds DNA with used the DNA as the carrier. The growth of Cu NCs were studied by transmission electron microscope(TEM) and fluorometry. The electrochemical behavior of Cu NCs were investigated by cyclic voltammetry(CV). The differential-pulse voltammetry(DPV) had also been used to investigate the analysis capability of the sensor by using Cu NCs as a response signal. The experimental results show that, the signal of Cu NCs was strengthened gradually with the increase of the target DNA concentration, and the oxidation peak current(Ipa) versus the logarithm of S2 concentration(lg CS2) exhibits a wide linear range from 1.0×10-14 mol/L to 1.0×10-20 mol/L with a detection limit of 1.01×10-21 mol/L. The selective binding experiment for the target DNA indicated that the biosensor also can well distinguish the target DNA from the single-base mismatch sequences, three base mismatch sequences and non-complementary sequences. At the same time, the proposed sensor also exhibited good stability, repeatability and reproducibility.(2) A Pb2+ electrochemical biosensor was constructed based on the selective adsorption of poly thymine(poly T) to Pb-Au alloys nanomaterials(Pb@Au NPs). The Au E were modified with DNA probes first,and then treated with MCH to block the non-specific sites on the surface. The Pb-Au alloy nanoparticles(Pb@Au NPs) with core-shell structure come from the specificity etching of gold nanoparticles(Au NPs) by sodium thiosulfate and Pb2+. Pb@Au NPs were assembled on the biosensor by the selective adsorption of poly thymine(poly T) to Pb@Au NPs, and electrochemical properties were tested. The experimental results show that, because of Pb@Au NPs was covered with thiosulfate which negatively charged, the impedance of [Fe(CN)6]3-/4- on the electrode surface increased after assembled Pb@Au NPs on the sensing interface. The reaction conditions such as DNA sequence, temperature and concentration of Au NPs, were investigated and optimized. Under the optimized conditions, the impedance values(Rct) versus the logarithm of Pb2+ concentration(lg CPb2+) exhibits a wide linear range from 1.0×10-15 mol/L to 1.0×10-10 mol/L with a detection limit of 3.22×10-16 mol/L. The proposed sensor shows excellent dual-specificity selectivity which based on the selective adsorption of poly T to Pb@Au NPs and the specificity etching of Au NPs by sodium thiosulfate and Pb2+.(3) A electrochemical Pb2+ biosensor was constructed based on in-situ growth of G-quadruplex templated copper nanoclusters(G4-templated Cu NCs) on the DNA modified interfaces. The Au E were modified with Pb2+ aptamer first,and then treated with MCH to block the non-specific sites on the surface. After Pb2+ induced Pb2+ aptamer fold into stable G-quadruplex structures, the electroactive G4-templated Cu NCs formed specifically by the disproportionation of Cu Br with G4 taken as the carrier. The growth of Cu NCs were studied by fluorometry, and the modification process of the sensor was characterized by EIS and CV. The DPV had also been used to investigate the analysis capability of the sensor by using Cu NCs as a response signal. The experimental results show that, the signal of Cu NCs was strengthened gradually with the increase of the Pb2+ concentration, and the Ipa versus the lg CPb2+ exhibits a wide linear range from 1.0×10-14 mol/L to 1.0×10-8 mol/L with a detection limit of 1.20×10-15 mol/L. At the same time, the proposed sensor also exhibited good selectivity, stability, repeatability and reproducibility.