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  Generic Process Scheme for Particle-Based Purification of Cell Culture Derived Influenza A Virus

Kalbfuss, B., Zimmermann, A., Genzel, Y., & Reichl, U. (2005). Generic Process Scheme for Particle-Based Purification of Cell Culture Derived Influenza A Virus. Poster presented at Bioperspectives, Wiesbaden, Germany.

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Item Permalink: http://hdl.handle.net/11858/00-001M-0000-0013-9C20-D Version Permalink: http://hdl.handle.net/11858/00-001M-0000-0025-20E1-A
Genre: Poster

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 Creators:
Kalbfuss, B.1, Author              
Zimmermann, A.1, Author              
Genzel, Y.1, Author              
Reichl, U.1, 2, Author              
Affiliations:
1Bioprocess Engineering, Max Planck Institute for Dynamics of Complex Technical Systems, Max Planck Society, ou_1738140              
2Otto-von-Guericke-Universität Magdeburg, ou_1738156              

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 Abstract: With the ongoing shift from traditional egg-based production of viral vaccines to cell culture-based cultivation, new purification strategies need to be developed to account for the different starting conditions. In the case of viral vaccines high loads of cellular debris and impurities have to be faced - in contrast to recombinant proteins or viral vectors produced by packaging cell lines. Here we present a modular process scheme for particle-based purification of human and equine influenza A virus as a model system. Virus was replicated with adherent Madin-Darby canine kidney cells, either in parallelized roller bottle cultures or microcarrier fermentations using serum-containing (fetal bovine serum) and serum-free media. Individual tasks were defined (clarification, inactivation, concentration, separation of impurities) and then rendered by appropriate unit operations. A traditional „harvest - capture - purification“ scheme was followed with the additional inactivation step inserted between the harvest and capture steps. Clarification was achieved by sequential depth and static membrane filtration with an option for centrifugal techniques in the future. The main purpose of this step was the removal of cell debris and gel particles contained in microcarrier cultures. Virus inactivation was conducted chemically with the well-established reagents b-propiolactone or ethylenimine. For the capture step, dynamic ultrafiltration, affinity chromatography and adsorptive filters have been investigated aiming at a first purification and concentration of the product. The capture step was then followed by a combination of chromatographic steps and/or adsorptive filters that were selected for efficient stripping of colloidal impurities, in particular host cell protein and dsDNA. In order to characterize each unit operation, robust microtiter plate assays have been developed for the quantitation of viral activity (infectious, hemagglutinin, neuraminidase), protein and dsDNA content. An immunological assay was employed to quantitate the virions. Particle size distributions were determined where appropriate using dynamic light scattering techniques.

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Language(s): eng - English
 Dates: 2005
 Publication Status: Not specified
 Pages: -
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 Rev. Method: -
 Identifiers: eDoc: 227070
 Degree: -

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Title: Bioperspectives
Place of Event: Wiesbaden, Germany
Start-/End Date: 2005-05-10 - 2005-05-12

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