Development Monoclonal Antibodies Against Ha Protein Of AIV H7 Subtype And NA Protein Of N1 Subtype And N9 Subtype

Avian influenza is an infectious disease of birds and poultry caused by RNA viruses of the family Orthomyxoviridae,the avian influenza viruses(AIV).Wild aquatic birds are considered natural hosts for the influenza a virus.According to antigenic variation in the major viral glycoproteins hemagglutinin and neuraminidase,the 16 hemagglutinin and 9 neuraminidase are perpetuated in aquatic birds.In recent years,the epidemic of AI represented by the H5N1 subtype and H7N9 subtype continues to cause a severe loss for poultry industry,and threaten both human health and public health security of the world.In this study,BALB/c mice were immunized alternatively with multiple branches of H7 avian influenza viruses that were isolated during the AIV epidemiological surveillance in our country.Five monoclonal antibodies(MAbs)against HA protein of H7 subtype,which named 4H4,4G8,4F7,4B8,3C1,were prepared by HI test and biological characteristics were analysed.4H4 was IgG2 a class and had a kappa light chain while the other MAbs were IgG1 class and had kappa light chains.HI test was performed with twenty three isolates which could be divided into three different clades.The result of HI test desmontrated that 4H4 MAb showed broad reactivity to H7 lineages besides of A/ENV/FJ/S3148/2013(H7N8).4H4 showed broad hemagglutination inhibiting.The other four MAbs only bound to part of H7 isolates.For most of H7 subtype AIV,they had not only a relatively conservative antigenic site,but a multiple of HA active antigenic sites.The results of neutralization test showed that five MAbs had broad neutralizing activity.IFA showed that five MAbs could bind to 293 T cell which was transfected into pCAGGS-H7.This study laid the material foundation for the identification and diagnosis of H7 AIV,and provided material means for studying the structure and function of HA protein.BALB/c mice were immunized alternately with pCAGGS-NA and viruses which have the same NA subtype and the different HA subtypes.Antibody isotypes were determined by IFA.Five hybridoma cell strains and seven hybridoma cell strains which could produce N1 MAbs and N9 MAbs repectively were developed and biological characteristics were analysed.These N1 MAbs were IgG2 b class and had kappa light chains,these N9 MAbs were IgG1 class and had kappa light chains.IFA showed that MAbs that we produced had cross reaction with some isolates.This study laid the material foundation for the method development of identification and diagnosis of NA antibodies of N1 and N9 subtype in serum.