Development Of A Immunochromatographic Electrochemical Assay For Cucumber Green Mottle Mosaic Virus

Cucumber green mottle mosaic virus(CGMMV)seriously harms cucumber,watermelon,bottle gourd,muskmelon and other cucurbit crops,and CGMMV will cause a large reduction in crop yield.In recent years,CGMMV has been detected many times in imported seeds by the inspection and quarantine departments of ports in China.The plants infected with CGMMV have different symptoms due to different environmental conditions,plant species and host plants.Generally,it will lead to plant growth retardation,dwarfism,mottled and deformed leaves,fibrosis and even rot in the flesh.It is necessary to establish a more sensitive,rapid and efficient detection method for CGMMV.Taking CGMMV as the research object,this paper optimizes the detection method of double-anti-sandwich immunochromatographic strip by using the existing monoclonal antibody(McAb)with high affinity and specificity in the laboratory,and preliminarily establishes the detection method of electrochemical analysis by immunochromatography,so as to realize the specific and rapid detection of CGMMV.The main contents include the following aspects:(1)Colloidal gold immunochromatographic strip detection methods of optimization:The lab secretion of prophase preparation can save the stability anti-CGMMV McAb hybridoma cell lines for recovery and subculture,preparation and collect the ascites,antibody.to make use of antibody affinity chromatography column purification,for subsequent related detection methods of optimization and building materials.The McAb 4D4 was used as the recognition antibody,it was labeled with colloidal gold,and McAb 2C9 was used as the capture antibody.Meanwhile,in order to improve the stability of the immunochromatographic strip,the parameters and working conditions of the strip were optimized,and the existing detection method of CGMMV immunochromatographic strip was improved and perfected.The optimized strip can be effectively used in CGMMV detection after being vacuum-sealed and stored at room temperature for 152 days,with good stability.By qualitative testing,it was determined that the detection limit dilution ratio of the test strip for the virus was 6400 times,and there was no cross-reaction with tobacco Mosaic virus(TMV)and cucumber Mosaic virus(CMV),which realized the rapid and specific detection of CGMMV.(2)Development of immunochromatographic electrochemical biosensor detection methods:This method is a rapid detection technology combining immunochromatography strip with electrochemical detection technology.In this chapter,anti-CGMMV McAb(4D4)and HRP labeled gold nanoparticles were used to synthesize dual-labeled nanoprobes(McAb-HRP-AuNPs)that can significantly amplify the detection signal,and the experimental parameters and working conditions of the electrochemical detection technology of immune chromatography were optimized.Immunochromatographic electrochemical biosensor technology combines immunochromatography strip of the rapid and electrochemical detection technology of high sensitivity,low energy requirements.test results,portable advantages of easy operation,intuitive easy to analysis for CGMMV preliminary build give a kind of new detection method,fast and efficient for the future to be able to develop more excellent detection technology.