Detection Of MC-LR Based On The Magnetic Beads And Aptamer Labeling Immunosensor

The structure of MCs is monocyclic heptapeptides,which could cause chronically poisons the liver and promotes liver cancer.Nowadays,more than 100 kinds of variants have been known already,and MC-LR is the most toxic and freshly harmed algae toxin in MCs.The World Health Organization has classified MC-LR as a neurotoxin and carcinogen.Traditional methods of detecting MCs generally require expensive equipment and cumbersome sample preparation.Thus,it was necessary to develop a fast,sensitive and reliable method to detect and quantify MCs in environmental samples owing to the strong toxicity and ubiquity of MCs.This not only improves the efficiency of testing,but also guarantees the safety of people's drinking water.Photochemical and electrochemical immunosensing are important means of rapid detection of microcystins.In recent years,nanotechnology has attracted the interest of many researchers,and the combination of nanotechnology and sensing technology has become a research hotspot of immunosensors.In this paper,three different photochemical sensors and electrochemical sensors were constructed using Fe₃O₄ nanobeads,metal nanoparticles and polydopamine nanospheres?PDA?to detect MC-LR in environmental water.In this project,we synthesized the biocompatible PDA and Fe₃O₄ nano-magneticbeads.PDA decorated with metal nanoparticles are used as a label,the functionalization of Fe3O4via coated with a layer of silica or polydopamine.The labels and magnetic beads are functionalized with aptamers and antibodies,and based on the antigens specific recognition with aptamers and antibodies,which the highly selective immunosensors could be constructed.In this dissertation,a series of research work which has been carried out is as follows:?1?The silica layer was coated on the surface of Fe₃O₄ nano-magneticbeads,which were used as the core,to form a surface-functionalized Fe₃O₄@SiO₂ core-shell structure.The functionalized Fe₃O₄@SiO₂ carboxyl group and the microcystins antibody?anti-MC-LR?were coupled to obtaine the Fe₃O₄@SiO₂-antibody complex.PDA/CuNPs complex was obtained by synthesizing PDA and then loading a amount of CuNPs on the surface.Then PDA/CuNPs complex was functionalized with carboxyl and coupled with MC-LR aptamer to obtain PDA/CuNPs-aptamer complex.In the presence of MC-LR,the MC-LR sandwich recognition method was used,and then the copper was dissolved and the color was developed to collect the ultraviolet absorption signal.After optimization of various conditions,the detection performance of the UV-visible immunosensor on MC-LR was investigated under optimal conditions.The method showed a good linear relationship to MC-LR with a detection range of 0.05-25.0 nM and a detection limit of 0.05 nM?S/N=3?.Furthmore,we demonstrated practical applicability of UV-visible sensor in detecting MC-LR in real sample with satisfactory results.?2?Fe₃O₄@PDA-antibody complex obtained by carboxyl-functionalized Fe₃O₄@PDA coupling with antibody as a material for separation and enrichment of immunomagnetic beads.PDA/CuNPs-aptamer obtained by coupling PDA/CuNPs with aptamers was used as labels to detect the signal response of MC-LR by indirect detection of copper ions fluorescent probe?PQ?.Various materials were characterized by Fourier transform infrared spectroscopy,X-ray diffraction,transmission electron microscopy,Raman spectroscopy,vibrating sample magnetometer,transmission electron microscopy,and nuclear magnetic resonance.A sandwich-type structure immunosensor was constructed by the dual recognition of the antibody and the aptamer in the presence of MC-LR.Finally,Cu²⁺acquired by the acid hydrolysis of CuNPs combines with the fluorescent probe?PQ?to generate a strong fluorescent signal,thereby achieving indirect detection of MC-LR.Under optimal conditions,the fluorescence signal response showed a good linear relationship with the concentration of MC-LR between 0.5 and 30.0 nM,and the detection limit was0.03 nM?S/N=3?.The fluorescence sensor also showed good sensitivity in the detection of actual water samples.?3?We explored an electrochemical linear scan stripping voltammetric signal based on silver nanoparticle functionalized PDA for the detection of MC-LR electrochemical immunosensors.The Fe3O4@PDA-antibody magnetic composite obtained by the coupling reaction of the antibody and the carboxyl functionalized Fe₃O₄@PDA used as a separation and enrichment material.Reduction of sodium citrate-stabilized AgNPs by NaBH₄ is negatively charged.The cationic polyelectrolyte of polydiallyldimethylammonium chloride?PDDA?combines AgNPs with polydopamine sphere by electrostatic interaction to obtain PDA/AgNPs as labels.Under optimal conditions,we investigate the linear range and detection limit of the electrochemical immunosensor for MC-LR using the direct immunization mode of antibody-MC-LR-aptamer.The detection range of the electrochemical sensor for MC-LR is 0.01-50.0 nM,and the detection limit is 0.001 nM?S/N=3?.We explore the practical application of the sensor and achieve satisfactory results.