Fabrication Of Novel Electrochemical Immunosensors And Its Application In Environment And Food Safety

Electrochemical immunosensor, which integrates the specific of immunoreaction between antigen-antibody with high sensitivity and diversity of electrochemical techniques, have been a promising analytical method since its advantages of simple operation, real-time output and ease of miniaturization. And these immunosensors have been widely used in cancer early diagnostics, environmental and food safety monitoring. Because the high cost and limited sensitivity to small molecule target of traditional immunoassay. this thesis has focused on fabrication of new type amperometric immunosensor combined with screen printing technology, supramolecular host-guest interactions and metal nanoparticles catalyst. The immunosensors possess potential application in fields of environmental public safety and food safety. The research of this work is as follows:1. Enzyme-free signal amplification for electrochemical detection of Mycobacterium lipoarabinomannan antibody on a disposable chipA simple, rapid, and disposable immunosensor at screen printed carbon electrode (SPCE) was developed by using gold nanoparticles (AuNPs) labeled Staphylococcal protein A (Au-SPA) as the electrochemical tag for detection of lipoarabinomannan antibody (anti-LAM). The immunosensor as the disposable chip was prepared by immobilizing capture antigen on screen printed carbon working electrode by passive adsorption, and characterized with scanning electron microscopy. After binding with the anti-LAM for further capture of Au-SPA. AuNPs was introduced as an electrochemical tag by the eletrooxidation of AuNPs in0.1M HC1to produce strong electroactive substance for signal amplification. Comparing with the enzyme-based immunosensor, AuNPs as enzyme-free tag for signal amplification exhibited many advantages such as no need of deoxygenation, and high stability. Under optimal detection conditions at preoxidation potential of+1.3V for30s, this method achieved the linear concentration of anti-LAM from15.6to1000ng mL-1with detection limit of5.3ng mL-1. The immunosensor showed a good performance with high selectivity, acceptable stability, and simple operation, providing a promising application as an adjunctive tool in early tuberculosis diagnosis.2. Host-Guest Interaction of Adamantine with p-cyclodextrin Functionalized AuPd Bimetallic Nanoprobe for Ultrasensitive Electrochemical Immunoassay of Small MoleculeA modular labeling strategy was presented for electrochemical immunoassay via supramolecular host-guest interaction between β-cyclodextrin (β-CD) and adamantine (ADA). The ADA labeled antibody (ADA-Ab) was synthesized via amide reaction, and the number of ADA moieties loaded on single antibody was calculated to be～7. The P-CD functionalized gold-palladium bimetallic nanoparticles (AuPd-CD) were synthesized in aqueous solution via metal-S chemistry, and characterized with transmission electron microscopy and X-ray photoelectron spectra. After the ADA-Ab was bound to antigen modified electrode surface with a competitive immunoreaction, AuPd-CD as signal tag was immobilized onto the immunosensor by the host-guest interaction, leading to large loading of AuPd nanoparticles. The highly efficient electrocatalysis of AuPd nanoparticles towards NaBH4oxidation produced an ultrasensitive response to chloramphenicol as a model of small molecule antigen. The immunoassay method showed a wide linear range from50pg/mL to50μg/mL and a detection limit of4.6pg/mL. The specific recognition of antigen to antibody resulted in good selectivity of the proposed method. The host-guest interaction strategy provided a universal labeling approach for ultrasensitive detection of small molecule targets.