Preparation Of High-Growth H1N1 Influenza Virus In Vero Cell By Reassortant Methods

Influenza is a kind of dangerous respiratory disease infected by influenza virus,which is characterlized by sudden outbreak and quick spread and can infect people and many kinds of animals and has trememendous effect to the world.Now the main method to prevent influenza is to inoculate vaccines.In the market the influenza vaccines are mainly the inactive vaccines made by chicken embryos. There are many aspects should be promoted of the influenza vaccine production by egg embryos.For example, the vaccine produced cope with influenza pandemic is not enough.Due to the disadvantages of chicken embryos,it is important to improve the influenza based on mammal cells. Vero cell is WHO resigned cell strain to produce vaccine and can be cultured at large scale by the bioreactor which has great foreground to produce influenza vaccine. But Vero cell is not the influenza virus susceptible cell naturally.So it is important to obtain high-growth H1N1 influenza virus in Vero cells for the influenza vaccine production by Vero cells.In this experiment we took Vero cell adapted influenza virus A/yunnan/1/2005Va (H3N2) as donor strain, inoculated together with the 2007-2008 vaccine virus strain A/Caledonia/ 20/99 (H1N1) into chicken embryos and Vero cells. The reassortant viruses were cultured and screened in Vero cells for two passages in the presence of antibodies specific to the donor virus.The screened viruses were continuously passaged for twelve times. The subtype was determined by HI test,Immunoflourescence Stain Assay,SRID. RT-PCR amplification of gene sequence analysis proved to contain A/Caledonia/20/99 (H1N1) of the HA and NA.The ICR mice were immunized with the inactivated influenza vaccine produced in Vero cells using the reassortant strain. The results showed reassortanted influenza virus(H1N1),which are adapted to Vero cells was prepared and there was no significant difference in immunogenicity of this strain while compared with parental strain A/Caledonia/20/99 (H1N1) .The conclusion is that the Vero cell adapted influenza virus A/yunnan/1/2005Va (H3N2) can be used as donor strain to prepare reassortmented vaccine strains adapted to Vero cells.