Efficient Production of Swine Influenza Virus Using Serum-Free Suspension MDCK Cells for Vaccine Manufacturing

Cell culture-based manufacturing platforms, already well-established for human influenza vaccines, are increasingly being adopted in the veterinary field as a powerful alternative to traditional embryonated hen's eggs, providing improved scalability and safety. In this study, we established a serum-free suspension culture process using Madin-Darby canine kidney (MDCK) cells for the efficient production of swine influenza virus (SIV). To optimize SIV production, we investigated multiple customized serum-free media in shaking flask cultures and selected an optimal formulation that supported efficient viral production. When this optimized medium was applied to a bioreactor culture, it supported robust cell growth, achieving a maximum viable cell density of 3.8 × 10⁶ cells/mL prior to infection. Subsequent direct SIV infection experiments, conducted without media exchange in both shaking flask and bioreactor cultures, consistently yielded high and stable virus titers reaching a maximum log₂ titer of 8.0, thus confirming the excellent scalability and reproducibility of the process. Moreover, the test trivalent vaccine developed in this study using the bioreactor-produced SIV antigen triggered high hemagglutination inhibition (HI) antibody titers and robust geometric mean titers (GMT) in animal models, thereby demonstrating superior immunogenicity to egg-based commercial vaccines. Taken together, these findings suggest that MDCK cell-based suspension culture platform may provide a scalable, high-yield, and reliable framework for the modern manufacturing of cell-based SIV vaccines.