Production of Novel Influenza A(H7N9) Virus Recombinant Ribonucleoproteins and Anti-Nucleoprotein Monoclonal Antibodies

In March 2013, the first human case infected by a novel avian influenza H7N9 virus subtype was reported in China. In April 2013, Taiwan CDC confirmed the first imported case of H7N9 infected patient from China. It is the first time that a low pathogenic avian influenza virus has led to human infection on account of genetic reassortment. Moreover, some isolated H7N9 viruses have shown Tamiflu drug-resistant. As of May, 2016, 786 documented human cases were confirmed by WHO and the mortality rate was as high as 39%. Therefore, it is urgently important to strengthen basic research of H7N9 and develop rapid laboratory diagnostics and antibodies. Influenza virus ribonucleoproteins (RNP), composed of NP, PA, PB1, PB2, and viral RNA, play important roles in viral gene transcription and replication. NP is a multifunction protein, which not only interacts with viral proteins but also interacts with host cell proteins. Therefore, NP is a potential antiviral target. The present study is aimed to express NP, PA, PB1 and PB2 recombinant proteins by for production of monoclonal antibodies. These four proteins have been stably generated in Sf21 insect cells and the NP recombinant proteins were purified by affinity chromatography. The anti-NP monoclonal antibody 2-4-7B showed great specificity and can be applied for NP immunoprecipition. In addition, this study found that the expression levels of PA, PB1 and PB2 proteins were increased by fusion of GP67 signal peptide at the protein N-terminus. However, PA, PB1 and PB2 were expressed as inclusion bodies, therefore the protein purification strategy should be further investigated.