Conversion of MDCK Cells to Suspension Culture and Preliminary Application in the Proliferation of Avian influenza virus H9 Subtype
LIU Ping1,2, CUI Yan1,*
1 College of Veterinary Medicine, Gansu Agricultural University, Lanzhou 730070, China; 2 China Agricultural Vet.Bio.Science and Technology Co., Ltd., Lanzhou 730046, China
Abstract The pandemic of avian influenza poses a serious threat to the breeding industry and public health which could cause great losses in national economy. Currently, vaccination against avian influenza is the most effective preventive measure. There are many problems in the traditional chicken embryo technology, which is gradually replaced by cell technology. In order to evaluate the application of suspension culture technology in the production of inactivated cell-derived Avian influenza virus (AIV) vaccine, a well-defined adherent canine kidney cells (MDCK) was selected and acclimated to the suspension condition by gradually reducing serum. The AIV H9 subtype was inoculated into the adherent and suspended MDCK for adaptation culture. The cell-derived and chicken embryo-derived virus were prepared into inactivated vaccine. The specific pathogen free (SPF) chickens (Gallus gallus) were immunized and the immune effect of cell-derived and chicken embryo-derived AI inactivated vaccine was evaluated by serological method. The results showed that the MDCK cell density proliferated to 3.0×106 cells/mL after 48 h of suspension culture, and reached to 5.0×106 cells/mL after 72 h with good morphology, high viability and single suspension in culture medium. The data and cell status of shaking flask and 7.5 L bioreactor culture showed that the cell strain was suitable for suspension culture in a bioreactor. The AIV H9 subtype proliferated by suspension MDCK cells was used to prepare inactivated vaccine. After 21 d of immunization with SPF chickens, the geometric mean value of Hemagglutinatio inhibion (HI) in serum was more than 6(log2), which was equivalent to the immune effect of chicken embryo-derived vaccine. The above experimental results suggest that the selected suspension culture MDCK cell line had the characteristics of rapid growth and stable proliferation, and could be used to proliferate AIV H9 subtype with high hemagglutination (HA) titer. The inactivated vaccine had good immunogenicity and could stimulate the body to produce antibodies with high titers. This study provides basic cytological data for large-scale production of AI and other viral vaccines.
LIU Ping,CUI Yan. Conversion of MDCK Cells to Suspension Culture and Preliminary Application in the Proliferation of Avian influenza virus H9 Subtype[J]. 农业生物技术学报, 2020, 28(1): 184-190.
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