Preparation And Identification Of Monoclonal Antibodies Against M Protein Of TGEV

E.coli BL21 harboring recombinant PGEX-6P-TM was induced with IPTG to express recombinant M protein. The BALB/c mice were immunized with purified rMP. Cells inosculation was carried out by standard method.The hybridoma supernatants were selected by indirect ELISA.The selected cells were subcloned three times by limiting dilution .Three hybridoma lines against TGEV rMP were obtained , which were designated as 1B3,3C2,4A5. The isotypes of the McAbs were IgG2a,IgG1,IgG1. The three hybridoma lines can secrete antibodies steadily after culturing and saving for a long time.The average numbers of chromosomes were all from 85 to 95 pairs. Their antibody titers of cells culture supernate were 1:4000,1:1000,1:4000 respectively. Their titers of ascites were 1:5×105,1:5×106,1:5×106 respectively.These McAbs didn't react with PEDV,PRV and PrV,but they were highly specific to TGEV by indirect ELISA.The result showed that these McAbs had high specificity. They can be used to diagnosis TGEV.These McAb were used to detect pathogeny by IFA. The result of IFA indicated that IBRS cells affected with TGEV showed strongly yellow and green fluorescent in cytoplasm and cell membranes after acting with McAbs,the cells inoculated TGEV had no strongly yellow and green fluorescence after acting with SP2/0 cells culture supernate. The McAbs against TGEV M protein were highly specific to detect pathogeny.All these results showed that the McAbs against TGEV M protein were highly specific to detect pathogeny.All the work founded material base for accurate and rapid diagnosis of TGEV.