Preparation And Preliminary Application Of Monoclonal Antibodies Against HA Protein Of AIV H3 Subtype

H3 subtype avian influenza virus is not as effective as highly pathogenic avian influenza virus in affecting poultry production,but in the process of virus transmission,H3 subtype influenza virus can provide gene fragments to other subtypes of influenza virus in poultry,thus strengthening the detection of H3 subtype influenza virus is very important.At present,there is no rapid detection method for H3 subtype avian influenza virus,to establish a rapid detection method against HA protein of H3 subtype avian influenza virus(AIV).DK/GX/S10225/2015(H3N2),DK/GZ/S1352/2015(H3N8),DK/FJ/S1131/2015(H3N3)was used as an immunogen to immunize six-week old BALB/c female mice.After three immunizations,mouse spleen cells were fused with SP2/0 cells under the action of PEG.Twenty-seven MAbs were screened by HI experiments and were named as 1E12,1F7,1-1C9,1-2G7,1-3C8,1-3G2,1-4B6,1-4E6,1-4E11,2F11,2-3G3,2-4B6,2-4F11,2-4H7,3B10,3C9,3-1A5,3-2A2,3-2D2,3-3F6,3-3G4,3-2H4,4A12,4H11,4-1B6,4-2F10 and 5-4D8 and biological characteristic were analysed.The heavy chain isotype of 1-3C8 belongs to IgM,the heavy chain isotype of 2-4F11,2-4H7,3-1A5,3-2A2 and 3-3F6 belong to IgGl,the heavy chain isotype of 1-1C9,1-3G2,1-4B6,1-4E6,2F11,3B10 and 3-2D2 belong to IgG2a,heavy chain isotype of 1E12,1F7,1-2G7,1-4E11,2-3G3,2-4B6,3C9,3-2H4,4H11 and 4-2F10 belong to IgG2b.The light chain isotype of twenty-seven McAbs are all the ?chain.HI results showed that 1-2G7,2-4H7,3-1A5,3-2A2 and 3-2H4 could cross-react with H6 subtype AIV,1-2G7,1-4B6,3-2A2 and 3-2H4 could cross-react with H11 subtype AIV,1E12,1F7,1-1C9,1-3C8,1-3G2,1-4E6,1-4E11,2F11,2-3G3,2-4B6,2-4F11,3B10,3C9,3-2D2,3-3F6,3-3G4,4A12,4H11,4-1B6,4-2F10 and 5-4D8 McAbs only could react with H3 subtype AIV,which have a high specificity.The 1-4E6,1-3G2,3-3G4,1E12,2F11 and 3-3F6 could react with H3 subtype AIV,and it has a broad spectrum.Western blot data were shown that the six McAbs all could bind to HA1 protein.Using the prepared broad-spectrum McAbs,the blocking ELISA method for detecting H3 antibody was established,which provided an effective tool for clinical diagnosis of H3 subtype avian influenza virus.