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Abstract

To increase process yields in virus vaccine production in mammalian cell culture high cell densities are of prior interest. As an example we investigate an influenza vaccine production process [1]. Madin Darby canine kidney cells (MDCK) were cultivated on Cytodex1 microcarriers and later infected with influenza A virus. To achieve maximum productivity in such a process one has to verify that specific virus productivities should remain unchanged with increasing cell densities. In high-density cell cultures process parameters such as medium and oxygen supply also have to be considered. Simpson et al. [2] reported for a hybridoma cell line in mAb production that the amount of apoptotic cells increased due to several stress factors, i.e. under elevated oxygen partial pressure.
Additionally, apoptosis is induced in the host cell during influenza virus infection (Takizawa et al. [3]). The virus induced apoptosis plays a crucial role during influenza virus infection as shown by Ludwig et al. [4]. Therefore, an investigation concerning the correlation between oxygen partial pressure-induced apoptosis and virus replication during vaccine manufacturing has been carried out. <p> Here, we present results obtained for high-density cell culture in small scale bioreactors. Cell numbers of 6.0 to 8.0 x 10 ⁶ cells/mL were achieved in cultivations with microcarrier concentrations of 10 g/L. Cultivations at 40 % of air saturation and above 150 % of air saturation will be compared with respect to cell numbers on microcarriers, virus titres, level of apoptosis (TUNEL assay), and glycosylation of viral haemagglutinin proteins (capillary gel electrophoresis–laser induced fluorescence (CGE - LIF)). First results indicate that apoptosis induced by elevated oxygen partial pressure can be beneficial for increasing virus yields. <p> [1] Genzel et al.; 2004. Vaccine 22: 2202-2208
[2] Simpson et al.; 1997. Biotechnology and Bioengineering 54: 1- 16
[3] Takizawa et al.; 1993. Journal of General Virology 74: 2347 - 2355
[4] Ludwig et al.; 2006. Cellular Microbiology 8(3): 375 - 386