The Spectral Analysis Of Biological Molecules Based On Gold Nanoparticles And Fluorescent Dyes

As an important nanomaterial,Gold nanoparticles(AuNPs) was widely applied in various fields by its unique physical and chemical properties. Aptamers are artificial single-stranded DNA or RNA sequences, show robust binding affinity for target to form the second or third structure formation which can improve the specificity of biosensor. Fluorescent dyes can be used to build the fluorescent signal which was related with the concentration of target. This thesis mainly used the spectral characteristics of Gold nanoparticles and fluorescent dyes to establish the different biological sensors for the detection of ractopamine, lysozyme, thrombin, mainly includes the following three parts:(1) A biological probe is based on gold nanoparticles and cationic fluorescent dyes for the detection of ractopamine. Gold nanoparticles were synthesized according to the previous report. Cationic fluorescent dyes caused the aggregation of gold nanoparticles through electrostatic adsorption leading to the fluorescence quenching of cationic fluorescent dyes. When adding ractopamine to the solution, ractopamine interacted with gold nanoparticles while the cationic fluorescent dyes were released. The recovery of fluorescence intensity for cationic fluorescent dyes was proportional to the concentration of ractopamine in the range of 0.91 to 273 nM, the detection limit was 0.28 nM. This method has the vital significance for quantitative detection of ractopamine in actual samples.(2) An aptasensor strategy was developed for detection of lysozyme based on gold nanoparticles and cationic fluorescent dyes. In the absence of lysozyme, the fluorescence of cationic fluorescent dyes was quenched when added to the aptamer and gold nanoparticles couldn't aggregate. The recovery of fluorescent related to the adding of lysozyme, the lysozyme hybridizes with its aptamer and release cationic fluorescent dyes. According to the above, the lysozyme can be detected simultaneously by using the colorimetric, UV-Vis and fluorescence methods. The experiment has good selectivity for detection of lysozyme with the linear range from 0.9 to 36 nM and the detection limit was 17 pM by fluorescence methods.(3) A label-free fluorescent strategy for thrombin detection based on exonuclease I(Exo I) and SYBR Green I assisted signal amplification has been demonstrated. In the absence of thrombin, the aptamer bound to its complementary DNA to form double-stranded DNA and the Exo I couldn't cause the splitting of ds-DNA. Once SGI dye was added, a weak fluorescence signal of SGI could be observed due to the intercalation of SGI into the grooves of the ds-DNA. In the presence of thrombin, the aptamer bound to thrombin and folded into a G-quadruplex structure, while which led to the dissociation of ds-DNA by Exo I and released SGI, more SGI intercalated into G-quadruplex structure and resulted in an enhancement of fluorescence intensity. This method provides a simple, sensitive, high specificity for detection of thrombin with the detection limit was 0.28 nM.