Nanoparticles And Small Moleculers Encapsulated In Nanocapsules For Fluorescent Immunoassay Of Pathogenic Bacteria

Food safety is directly related to the healthy development of human beings.Foodbome pathogenic bacteria have brought a great threat to human health.Escherichia coli 0157:H7 is one of the most common foodbome pathogenic bacteria.The infectious dose of Escherichia coli O157 H7 is less than 10 cells.To reduce the infection of pathogens and enable effective clinical diagnosis,it is urgent to develop a fast,sensitive and reliable method for the detection of Escherichia coli 0157:H7.In this thesis,Several rapid,sensitive and reliable methods for detecting Escherichia coli 0157:H7 are.The main contents are as follows:?1?Au nanoclusters?AuNCs?-embedded chitosan?CS?nanocapsules were prepared and used as a novel signal amplification system for sensitive fluorescent immunoassay of Escherichia coli 0157:H7.AuNCs@CS nanocapsules were prepared by pH-triggered self-aggregation of CS in the presence of AuNCs,where a large number of AuNCs were embedded in each nanocapsule.A rapid,ultrasensitive,and selective method for fluorescent detection of Escherichia coli 0157:H7 was developed by coupling the nanocapsule-amplification strategy with immunological recognition and magnetic separation.Compared with a conventional immunoassayusing AuNCs as labels,the fluorescent signals of the proposed immunoassay were greatly amplified,because much more fluorescent AuNCs are linked to each bacterial cell.As a result,a linear range was obtained from 3 to 700 CFU mL^-1,with a detection limit of 1 CFU mL^-1.This method was successfully used to detect Escherichia coli O157:H7 in drinking water and milk samples.?2?L-cysteine?Cys?-encapsulated liposom enanocapsules were prepared and used as novel labels for sensitive immunoassay of Escherichia coli O157:H7.Experimentally,antibodies modified nanocapsules and antibodies modified Fe3O4 nanoparticles can specifically bind to Escherichia coli O157:H7.After mangnetic seperation,nanocapsules linked to bacterial cells were collected,and then the nanocapsules were destructed by Tween-20 and Cys can be realesed from the nanocapsules.These Cys moleculers released from nanocapsules can react with colorless 4-chloro-7-nitro-2,1,3-benzo oxoxy heterooxazole?NBD-Cl?and thus produce yellow NBD-Cys molecules.Because NBD-Cys can produce yellow-greenfluorescence,a "OFF-ON" fluorescent method for the detection of Escherichia coli O157:H7 was developed.In addition,yellow NBD-Cys molecules were produced after the reaction of colorless Cys and colorless NBD-Cl,enabling colorimetric detection of Escherichia coli O157:H7.The linear range for fluorescentdetection of Escherichia coli O157:H7was from 4 to 3000 CFU mL^-1,with a detection limit of 1 CFU mL^-1,while the linear range for colorimetric detection of Escherichia coli O157:H7 was from 40 to 3000 CFU mL^-1,with a detection limit of 10CFU mL^-1.?3?Au nanoclusters?AuNCs?-embedded organosillica nanocapsules?BONs?were prepared and used as labels to constuct a sandwich-type immunosensor for fluorescent and electrochemical detection of Escherichia coli O157:H7.Electrochemical impedance spectroscopy was used for electrochemical detection of Escherichia coli O157:H7.AuNCs@BONs linked to bacterial cells were destructed by NaBH4 reduction,and fluorescentAuNCs can be released and used for fluorescent detection of Escherichia coli O157:H7.The proposed fluorescent and electrochemical dual-signal method can improve the accurancy for the detection of Escherichia coli O157:H7.The linear range for fluorescent detection of Escherichia coli O157.H7 was from 100 to 107 CFU mL^-1,with a detection limit of 30 CFU mL^-1,while the linear range for electrochemicaldetection of Escherichia coli O157:H7 was from 200 to 107 CFU mL^-1,with a detection limit of 100 CFU mL^-1.