Production And Characterization Of Monoclonal Antibodies Against Asia1 FMDV And The Development Of An Antigen Capture Elisa For The Detection Of FMDV

A panel of monoclonal antibodies (mAbs) against FMDV Asia1 , named as 1B4,1F5,3D9,3E11,3H6 and 5G3 ,were produced by fusing between SP2/0 myeloma cells and spleen cells of Balb/c mice immunized with the purified virus preparations. The antibody titers in cell supernatant and ascites were 1:1024,1:256,1:256,1:256,1:64,1:128 and 5×10-4,1×10-4,1×10-4,1×10-4,1×10-4,8×10-3, respectively. Enzyme-linked immunsorbent assay (ELISA) and indirect immunofluorescence assay(IFA) showed that mAbs 1B4,1F5,3E11,3H6 and 5G3 reacted with Asia1 FMDV, but not with O FMDV , revealing that they are specific to Asia1 FMDV. While mAb 3D9 reacted with both Asia1 FMDV and O FMDV,suggesting that its binding epitope is conserved between these two serotypes. Western blot analysis suggested that the epitopes they recognized were conformational since they failed to reacted with any viral proteins. The result of additive ELISA showed that they could recognize different antigen epitopes. The reactivity of these mAbs against trypsin-treated FMDV was examined using an indirect ELISA. The binding sites of the mAbs, are trypsin sensitive. Their relative affinity index (RAI) were 1.5mol/L,2.5mol/L,3.5mol/L,2.5mol/L,1.0 mol/L and 1.5 mol/L, which were indicated by thiocyanate elution measurement.An antigen capture ELISA for the detection of Asia1 FMDV was developed by using bovine polyclonal antiserum as capture antibody and monoclonal antibody 1B4 as detector antibody. With this assay the purified Asia1 FMDV was detected at a concentration of 109ng/100μL, and the antigen in the infected BHK-21 cells could be detected when the FMDV titer was 103 TCID50 /100μL. No false positive results were given when 10 negative samples were tested, and no cross-reaction occurred when O FMDV, contagious bovine pleuropneumonia antigen, bovine tuberculosis antigen, infectious bovine rhinotracheitis virus and bovine ephemeral fever virus were tested.In conclusion, these mAbs would be useful tools in the future studies of Asia1 FMDV. The establishment of the monoclonal antibody-based antigen capture ELISA would provide valuable experience for the development of diagnostic kits for the detection of FMDV.