Detection Methods Of Materials Which Play Important Roles In The Process Of Life Activity

The detection of adenosine, nucleic acid and hydrogen peroxide is of great importance to human being due to their significant roles in the process of life activity. The existing detecting approaches generally include interface detecting system and homogeneous detecting system. In present works, we have developed electrochemical biosensors and colorimetric detection techniques based on the latest research achievement of nanomaterials and aptamer, and obtain excellent characteristics, such as simplicity, high sensitivity, veracity and low-cost. These research works make positive contribution to the development and innovation of the analysis methods.1.Aptamers are nucleic acids that have high affinity and selectivity for their target molecules. A target may induce the structure switching from a DNA/DNA duplex to a DNA/target complex. In chapter 2, a reusable electrochemical sensing platform based on structure-switching signaling aptamers for highly sensitive detection of adenosine is developed. A gold electrode is first modified with polytyramine and gold nanoparticles. Then, thiolated capture probe is assembled onto the modified electrode surface via sulfur-gold affinity. Ferrocene (Fc)-labeled aptamer probe, which is designed to hybridize with capture DNA sequence and specifically recognize adenosine, is immobilized on the electrode surface by hybridization reaction. The introduction of adenosine triggers structure switching of the aptamer. As a result, Fc-labeled aptamer probe is forced to dissociate from the sensing interface, resulting in a decrease in redox current. The decrement of peak current is proportional to the amount of adenosine. The present sensing system could provide both a wide linear dynamic range and a low detection limit. In addition, high selectivity, good reproducibility, stability, and reusability are achieved. The recovery test demonstrates the feasibility of the designed sensing system for an adenosine assay.2. Guanine-rich DNA sequences commonly form helical quadruplex structures via Hoogsteen hydrogen bonds. The aggregation behavior of the nanoparticles, which are functionalized with four-guanine-terminated 27-base sequences at a nanoparticle- to-DNA ratio of 1:60, is investigated in chapter 3. To some extent, the guanine- quadruplex structures between the gold nanoparticles (GNPs) promote nanoparticle aggregation. However, the coordination site of the metal ion on the nanoparticle surface is partially passivated: the stability of guanine-rich DNA-GNPs is slightly lower than that of the usual DNA-GNPs, and the metal-ion specificity of nanoparticle assembly is substantially decreased. Thus, a mechanism for the aggregation of guanine-rich sequence-modified GNPs is proposed. It is possible to obtain a stable guanine-rich sequence-functionalized nanoparticle solution at high ionic strength by regulating guanine-rich DNA sequences. The controllability of guanine-rich sequence-modified nanoparticles makes the secondary structure of DNA a potentially useful candidate for DNA analysis and disease diagnostics.3. An exciting discovery that the adsorption of adenosine-5'-monophosphate (AMP), a negatively charged molecule, can induce the aggregation of gold nanoparticles has been reported in chapter 4. Along this line, a homogeneous, colorimetric system for nuclease characterization is described based on the intensive aggregation of unmodified gold nanoparticles induced by cleavage products using S1 nuclease as a model molecule. The present method exhibits the simultaneous advantages of simplicity, rapidity, reliability and low cost as covalently attached labels, the third DNA sequence or additional signal amplification is not involved. If this technique is used for the detection of DNA sequences, excellent response characteristics (for example, sensitivity, selectivity and linear response range) are achieved using either UV/vis spectrophotometer or the naked eye. The proposed technique associated with the adsorption of cleavage products onto gold nanoparticles opens a new opportunity for nuclease characterization as well as for genetic analysis.4. Based on the excellent features of gold nanoparticles such as large surface area, high catalysis effect, strong adsorption ability, nice biocompatibility and conductive ability,a mediator-free hydrogen peroxide (H2O2) biosensor has been developed in chapter 5. First, HRP was adsorbed by gold nanoparticles, then, it was immobilized on the bare gold electrode surface by activated concanavalin A (Con A). The experimental conditions were optimized. The electrode provided a linear response to H2O2 over a concentration range of 5.0×10-6-1.2×10-2 M with a detection limit of 2.9×10-6 M. The system was applied to the determination of samples, the results were satisfactory.