Preparation And Preliminary Application Of Monoclonal Antibody Against Japanese Encephalitis Virus

Encephalitis also known as Japanese encephalitis,is an important mosquito-borne zoonotic disease that caused by JEV.Pig is the main reservoir host to JEV.Encephalitis can seriously affect the survival rate of piglet,slaughter pig fattening and breeding pig in the litter,bringing huge losses to the pig industry.Meanwhile,JE is also a serious threat to human health.Therefore,to explore the diagnosis and treatment of encephalitis can lead an important significance to control the spread of JEV.In this study,use BHK-21 cell to Proliferation JE SA-14-14-2 virus strain,purified JEV were used to immune Balb/c mice,fusion spleen cell with SP2/0 cell,the indirect ELISA method was used to preliminary screening,by 3 times limiting dilution cloning,eventually obtained two cell strains that can secrete anti-JEV monoclonal antibody,named as B10 and C4.Analysis of monoclonal antibody and hybridomas cell biological characteristics,including subtype,titer determination,affinity analysis,cross-experiment,karyotype and stability of antibody secretion.The results showed that:McAb of B10 and C4 are all belong to lgG1subtype;titer of B10 McAb was 1:10 0000, titer of C4 McAb was 1:240000;the affinity were C4 McAb higher than B10 McAb;cross-experiment demonstrate that antibody of the two cell strains secretion were specificity;hybridoma cell with the treatment of successive passage and cryopreservation, the results showed hybridoma cell can stable secreting antibody;the chromosome number of B10 and C4 hybridoma were between 90-100.Use purified monoclonal antibody to establish double antibody sandwich ELISA to detection of JEV positive serum,the experiment result showed that the method has good sensitivity,can be better used for the detection of JEV,and provides a alternate means for the detection of JEV,and the method have a good prospect.