Preparation Of Monoclonal Antibodies Against Hemagglutinin Of Subtype H5 Avian Influenza Virus And Establishment Of Capture ELISA

Avian influenza is an infection and disease syndrome of birds (poultry and wildfowl) caused by influenza type A virus. The disease broke out in many countries of the world, and resulted in a serious economic loss. At the same time, it threatens the public health. Thus, it has a great practical importance to study the disease.H5 subtype avian influenza virus A/Chicken/Guangdong SZ/l/00(H5Nl) (SZ) strain was propagated in the allantotic cavity of 9¹1 day-old embryonated SPF chicken eggs. The allantotic fluid infected SZ was purified, and inoculated to 6⁸ week-old BALB/C mice. By using the hybridoma technique, SP2/0 myelomacells and spleen cells of the immunized BALB/C mice were fused in vitro. The two monoclonal antibodies (McAbs) were obtained by HI test and named 1E5, 4A4 alone. The immunoglobulin subtypes of the two McAbs were identified as IgG1, Kappa light chain. The HI titers of culture supernatants and ascites were 2⁶²⁸ and 2¹⁰²¹⁶ respectively. The two McAbs reacted specifically to AIV H5, but not to AIV H9, AIV H7, NDV and EDSV. The 4A4 McAb only reacted to AIV H5 hemagglutinin protein band by Western-blot.Capture ELISA technique for detecting H5 avian influenza was established with biotin labeled McAb. The results showed that the ELISA could detect H5N3 reference strain and all the H5 AIV isolates, but not react to the other H9 and H7 AIV isolates, NDV, IBDV and EDSV. So, the method has a wonderful application future in fast detection of AIV H5.