Yvonne Genzel

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..Here, glutamate and aspartate increase correlated to virus release kinetics, indicating cell disruption and apoptosis. Starting with a moi of 0.025 resulted in a maximum virus yield of 2.4 log HA/100 microl at 44 h post-infection...

..6 microM and quantitation limits of less than 1.6 microM. As an example, data for the amino acid composition of different media used for the production of inactivated influenza vaccines in cell culture are shown...

..After virus infection completely different profiles were found for essential and nonessential amino acids...

..Omission of the medium exchange before infection clearly simplified the process and reduced sterility risks due to washing steps...

..However, using these cultivation conditions in lab-scale bioreactors (stirred tank, wave bioreactor) resulted in lower productivities for Vero cells...

..These encouraging developments for cell culture-based influenza virus production are summarized and an overview on potential cell substrates is given...

..A detailed metabolic characterization of an influenza infected adherent cell line (MDCK) was carried out based on extracellular and intracellular measurements of metabolite concentrations...

..The findings will improve understanding of host cell response mechanisms during influenza vaccine production and viral strategies to evade these responses and to replicate efficiently in different cell lines...

..In Vero cells a selection for "rescue" variants was characteristic. After back-adaptation to MDCK cells some variants persisted at indifferent frequencies, others slowly diminished and even dropped below the detection limit...

..Absolute values of intracellular metabolites extracted with this modified method are comparable to the results of the two previously optimized methods, indicating a good extraction procedure according to the chosen response variables...

..From the results it can be concluded that the suggested process scheme is well suited for the harvesting and concentration of cell culture-derived influenza A virus...

..Comparing this minimum substrate consumption flux distribution to the fluxes estimated from experiments unveiled high overflow metabolism under the applied process conditions...

..Thus, the switch from glutamine-containing to glutamine-free medium with pyruvate provided a series of benefits without dramatic changes of cellular metabolism...

..Hence, altered lectin specificities have been observed between the two cell lines, requiring process adaptations between different influenza vaccine production systems...

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..Quantitative proteome-wide profiling of virus infection can provide insights into complexity and dynamics of virus-host cell interactions and may accelerate antiviral research and support optimization of vaccine manufacturing processes...

..9 hpi viral genome replication could be detected. The vRNA(-)s were synthesized in equimolar amounts and similar dynamics whereas preferential synthesis of NS1 vmRNA(+) in early transcription phases and a delay for M1 vmRNA(+) was found...

..In contrast to this, a principal switch of the N-glycan type attached to HA was observed when comparing different virus types (A and B) and subtypes (H1N1 and H3N2)...

..Flow cytometry represents a valuable tool to investigate the influences of cultivation conditions and process variations on virus replication and virus yields...

..abstract:..

..99 for most analytes. Working ranges were between 0.125-3.875 and 4.5-139.5 microM. Sample pH had a major impact on the quantification of several metabolites, but measurements were robust within a pH range of 6.5-9.0...