June 24, 2013

Scale-up of adherent Vero cells grown on Cytodex microcarrierrs using ReadyToProcess equipment


In cell culture-based vaccine production, scale-up of adherent cells is challenging. This study shows a process for scaling up adherent Vero cells from static cell factories to influenza production at 50 L scale using WAVE Bioreactor™ systems and ReadyToProcess singleuse equipment. Vero cells were grown to high cell density on Cytodex microcarriers in 10 L working volume. The cells were detached with trypsin and used to seed a 50 L production culture with the same microcarrier concentration. The cells were allowed to reattach and grow on the new microcarriers in a larger Cellbag™  bioreactor chamber. Cells were subsequently infected with influenza virus. The results show a repeatable scaleup procedure. The cell growth was similar in the 10 L seed cultures as in the 50 L production cultures in three consecutive experiments, with an obtained Hemagglutinin (HA) concentration of approximately 12 μg/mL and a virus titer of 109 virus particles/mL.


Introduction
Compared to an egg-based vaccine production, cell culture based vaccine production is seen as an alternative way for a more rapid response to pandemic challenges. Traditionally, cell-based vaccine production is performed in stainless steel bioreactors requiring extensive cleaning procedures. Withdisposable bioreactor systems without the need for cleaning and validation, the start-up time for GMP manufacturing can be significantly reduced (1). 

 
Adherent cells, such as Vero cells, are almost exclusively used for human cell-based vaccine production. Vero cells are susceptible to a broad range of viruses and are widely used for the production of viral vaccines like polio, rabies, or enterovirus (2, 3). Recently, influenza vaccine was successfully produced at industrial scale using Vero cells (4). Vero cells can only proliferate when provided with a suitable surface. 

 
The scale-up of such cell cultures is challenging because of the required detachment and reattachment of cells during passaging. The aim of this study is to present a process for scaling up adherent Vero cells for influenza propagation. The cultures were run at a working volume of 50 L using WAVE Bioreactor system and ReadyToProcess single-use equipment (Fig 1). When scaling up to 50 L from a 10 L bioreactor culture, the cells were detached by trypsinization and subsequently used to seed new Cytodex microcarriers of the same concentration in the larger Cellbag bioreactor. 

 
The cells reattached and grew on the new microcarriers and were subsequently infected with influenza virus. Our results show that the scale-up procedure using single-use WAVE Bioreactor systems is a fast and convenient option for virus production.



To see an overview of a cell-based influenza virus production process using single-use equipment, see this white paper: “Overview of a scale-up of a cell-based influenza virus production process using ReadyToProcess™ equipment”

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Tags: vaccine; influenza; readytoprocess; single-use; cytodex; microcarriers; cell-culture; wave; bioreactor