Stability of Minimum Essential Medium functionality despite L-glutamine decomposition.

L-Glutamine (L-Gln) instability in liquid media is a well-known fact. Also, negative effect of ammonia, one of the L-Gln degradation products, on viability of many cell cultures and on replication of different viruses has been described. However, negative effects of ammonia have been reported in doses excessively exceeding those that could be generated in regularly used liquid culture media due to spontaneous L-Gln breakdown (below 2 mM). Traditional virus vaccine production processes have been established and registered involving L-Gln containing media use. Eventual culture media replacement in the regular production process belongs to the major regulative changes that require substantial financial expenses. The aim of this study was to evaluate the effect of storage of Minimum Essential Media with Hanks salts on their relevant biological functions during virus vaccine production process in relation to L-Gln decrease. Our results show a cell type dependent effect of spontaneous L-Gln degradation during medium storage. They also suggest that for cell cultures used in measles, mumps, and rubella virus production the media retain their functionality in respect to cell viability or virus growth over a certain time window despite L-Gln degradation.

Critical factors for the replication of mumps virus in primary chicken embryo fibroblasts defined by the use of design of experiments (DoE).

Live attenuated vaccines against mumps virus (MuV) have been traditionally produced by passaging the virus in the embryonated chicken eggs or primary chicken embryo fibroblasts (CEFs). Virus propagation on these cell substrates enables successful virus attenuation and retains it sufficiently antigenic to induce lasting protective immunity in humans. The aim of this study was to identify critical factors for MuV replication in primary CEFs grown on a small-scale level in order to explore possibilities for improvements in the virus replication and yield. The effect of differently prepared cells, culturing conditions, and infection conditions on virus yield was estimated by employing statistical design of experiments (DoE) methodology. Our results show that the preparation of primary CEFs and the way of their infection substantially impact virus yield and are critical for efficient MuV replication. These process parameters should be considered in further process optimization. We also demonstrate the applicability of DoE in optimization of virus replication as a crucial step in obtaining high virus yields.