The Application Of Novel Sandwich Assay In Surface Plasmon Resonance Sensor

Surface plasmon resonance?SPR?biosensor is an optical biochemical sensor.The analytical principle of SPR biosensor is to study the biomolecular interaction and determine the analyte concentration accurately by collecting and analyzing the change of the medium refractive index which is caused by the change of medium properties or mass on the surface of the metal film.SPR biosensor has advantages of real-time monitoring,no marking and less sample consumption,and consequently shows an excellent prospect in immunological detection,medical diagnosis,food analysis and environmental monitoring.However,due to the limitation of testing methods and the structure of SPR biosensors,the detection sensitivity of the analyte at low concentration cannot meet current requirements,which limits the further development of SPR sensors.Generally,adjusting the properties of SPR sensing films and some indirect methods have been adopted to improve the detection sensitivity.Noble metal nanoparticles,especially gold and silver nanoparticles,are commonly used to improve the properties of SPR biosensor films for their superior optical characteristics.In addition,the sandwich assay is often used to improve the sensitivity and specificity of SPR biosensors by introducing secondary antibodies to increase the volume and mass of the complex molecules on sensor films.In this paper,gold,silver and other metal nanoparticles are combined with the sandwich assay and applied in SPR biosensors.Through the specific reaction,we can effectively increase the amount of adhesion on the sensing film surface,thereby significantly improving the performances of the biosensor,and then obtain an enhanced SPR biosensor.The main contents of this article are shown below:1.A novel SPR biosensor,which is based on hollow gold nanoparticles?HGNPs?and sandwich assay,was constructed and used to detect cardiac troponin I?cTrop I?.The gold sensing film was modified by HGNPs and polydopamine?PDA?through the immersion method.In order to amplify the SPR response signal,the sandwich assay was introduced.First,antibodies were connected with PDA to fix them on the surface of the sensor film.Then antigens were combined with the antibodies by the specific reaction.After that,secondary antibodies were directly modified on to the antigens to form a sandwich structure.The response concentration range of the new SPR biosensor for cTropI was 0.0375-40.00?g m L^-1,and the detection limit was 67times lower than that of the traditional SPR biosensor based on MPA.2.A new sandwich assay was applied to the SPR biosensor,using the secondary antibody-polydopamine-Ag@Fe₃O₄/reducedgrapheneoxide?Ab₂-PDA-Ag@Fe₃O₄/rGO?as a signal amplification agent.Ag@Fe₃O₄/rGO was prepared by a one-pot method and covered with PDA to form PDA-Ag@Fe₃O₄/rGO.Afterwardssecondaryantibodies?Ab₂?wereconnectedtoform Ab₂-PDA-Ag@Fe₃O₄/rGO.In the process of testing,the antibody?Ab₁?was first attached to the HGNPs modified gold film,and then specifically bound to the sample antigen.Finally,Ab₂-PDA-Ag@Fe₃O₄/rGO was introduced into the detection system to obtain the large sandwich structure as Ab₁/antigen/Ab₂-PDA-Ag@Fe₃O₄/rGO.The detection limit of the new SPR biosensor for rabbit IgG was 0.019?g mL^-1,which is132 times lower than that of traditional SPR biosensor modified with MPA,and 16times lower than that of HGNPs modified gold film biosensor.