COMMERCIAL EVIDENCE
Intensification of a PER.C6®-based recombinant Ad35 manufacturing process to prepare for commercial scale production
Marcel de Vocht
Crucell, NL 2008
This presentation shows the rapid progress Crucell is making with their TB vaccine manufacturing process. The ATF System is used in a variety of different ways at Crucell, for example cell banking and a modified perfusion application. They have reported such good gains in productivity that, coupled with disposable bioreactors, they have been able to avoid building a new facility for their manufacturing, and expect to produce the market supply needed with a modest retrofit of their existing facility.
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Continuous Perfusion Process in a Disposable Stirred-tank Bioreactor: A Case Study
Barbara Chiang, Nicole Richardson, Hong Haddock, Christian Wood, Ravi Bhatia and Sadettin Ozturk
Centocor R&D, USA 2007
Abstract: Disposable bioreactor systems are making their way into production of therapeutic products. A 28-day perfusion run was performed in the disposable stirred-tank using an external cell retention device to retain cells in the bioreactor. Results will show that the disposable stirred-tank was able to sustain high cell densities (> 15x106 cells/ml) and productivity comparable to conventional stainless steel bioreactor. Also, in this case study we will present equipment design challenges to scale-up perfusion process in the disposable bioreactor system.
Refine Edit: 15m cells is no longer considered high, of course!
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Equipment design and process challenges with running a perfusion process in a disposable stirred-tank: A case study
Nicole E. Richardson, Associate Engineer; Barbara Chiang, Assistant Engineer
Centocor R&D, USA 2007
A presentation similar to the poster above, but with more detailed process and engineering data shown. Some scale-up challenges are also discussed.
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Developments in cell culture perfusion processes and their scale-up to manufacturing
John Bonham-Carter
Magellan Instruments, UK 2005
This is an early presentation made to the UK branch of ESACT on data from the ATF System at a time when cell concentrations of 20m/ml was thought to be high: less than 5 years later, cell concentrations 10 times higher are being routinely achieved! It is also the first time that a concentrated fed-batch operation using ultrafiltration hollow fibre modules was presented, and the first time an explanation of how a reduction in operating costs for clarification can be achieved.
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Continuous and Scalable Production from a Bioreactor
Jim Furey
Refine Technology, USA 2002
Abstract: Batch, fed-batch, and perfusion are some of the typically used modes of operation for a cell culture based production process. Using process economic modeling and considering the scalable nature of the alternating tangential flow (ATF) filtration perfusion process; the most effective process mode to meet production requirements may be determined.
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ACADEMIC PUBLICATIONS
Fed-batch or perfusion for the production of biopharmaceuticals by animal cell cultivation?
Véronique Chotteau, Ph.D.
Bioprocessing Group, Royal Stockholm Technical University (KTH), SE 2009
KTH has performed an initial series of experiments with the ATF System to test its application in cell culture with a low performing cell line. Results, simple economic models and a comparison of other available perfusion systems are presented.
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Study of a perfusion process of Chinese Hamster Ovary cells by ATF filtration in bioreactor
Karin Tördahl, Philip Perroud, Véronique Chotteau, Ph.D.
Bioprocessing Group, Royal Stockholm Technical University (KTH), SE 2009
A second series of experiments was carried out and they are described here, covering high cell density cultures and the problems this brings, such as oxygen transfer limitation.
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(Poster) Study of a perfusion process of Chinese Hamster Ovary cells by ATF filtration in bioreactor
Véronique Chotteau, Karin Tördahl and Philip Perroud
Bioprocessing Group, Royal Stockholm Technical University (KTH), SE 2009
Abstract: Perfusion is a mode of operation where a continuous replacement of the conditioned medium by fresh medium is operated. It has the advantage of allowing high cell densities. This mode of operations is also required for some instable proteins since the cell-free supernatant containing the product of interest is immediately stored at low temperature where the proteolysis is not active. The ATF filtration device, Alternating tangential flow, has been designed to perfuse mammalian cell cultivation process. The cell broth circulation back and forward in the filter prevents the filter clogging and the design ensures a low shear not damageable for the cells. The purpose of the present work was to develop and study a perfusion process of Chinese Hamster Ovary cells producing a monoclonal antibody by ATF filtration in a 2 L working volume bioreactor. A serum-free medium was used (Irvine Scientific IS CHO CD XP). Cell densities above 40 x 106 cells/mL were obtained, up to 48 x 106 cells/mL. These high cell densities were challenging for the aeration. Pure oxygen aeration by large bubbles from an open tube resulted in satisfying oxygenation until 25 to 30 x 106 cells/mL but became limiting at higher cell densities due to the low kLa of these bubbles and the small liquid height. At higher cell densities, a porous sparger with pure oxygen was used either alone or in combination with the open tube aeration. PDF of this poster attached.
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Production and Glycosylation Analysis of Model Proteins from a Vaccinia Virus-Mammalian Cell Expression System
Nicole A. Bleckwenna,b,c, William Bentleyb,c, and Joseph Shiloacha
USA, 2003
a Biotechnology Unit, NIDDK, National Institutes of Health, DHHS, Bethesda, MD
b Center for Biosystems Research, UMBI, College Park, MD
c Department of Chemical Engineering, UMCP, College Park, MD
Abstract: A vaccinia virus-mammalian cell expression system was developed as an alternative method for recombinant protein production utilizing EGFP as a reporter protein (1). In previous work, EGFP production was evaluated in T-flask culture and in both suspension and microcarrier based bioreactor systems, where general production parameters were defined. In this work, the production capability of the system, as defined with EGFP, was evaluated using two proteins, the HIV gp120 envelope glycoprotein and hGC-1 (2), an olfactomedin-related protein. These proteins contain complex post-translational modifications, required for gp120 activity and possibly for hGC-1, although there is little information on this recently discovered protein. Two recombinant vaccinia virus strains were engineered with the genes for gp120 or hGC-1 and expression of these proteins was achieved in either T-flask culture or both T-flask and bioreactor culture by infection of the cell culture with recombinant virus. The production process, purification protocol and glycosylation pattern of gp120 is described. Bioreactor culture produced secreted gp120 up to 40 mg/L at 66 hours post infection (hpi).
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Scaleable Protein Production in Anchorage Dependent Mammalian Cells
Nicole A. Bleckwenna,b,c, William Bentleyb,c, and Joseph Shiloacha
USA, 2003
a Biotechnology Unit, NIDDK, National Institutes of Health, DHHS, Bethesda, MD
b Center for Biosystems Research, UMBI, College Park, MD
c Department of Chemical Engineering, UMCP, College Park, MD
Abstract: Recombinant protein production is a common method for providing clinical and commercial quantities of human therapeutic agents. Methods of production include recombinant bacterial fermentations, transfected or transformed cell culture, and virally infected cell culture. A potential protein production method with anchorage dependent mammalian cells has been evaluated. The system utilizes a recombinant vaccinia virus with the VOTE expression system that can potentially be used to produce large, highly glycosylated recombinant proteins, which may be difficult to produce by other means. A reporter protein, enhanced green fluorescent protein (EGFP), has been used to study the characteristics of this protein production method. Growth, infection and production parameters such as multiplicity of infection, culture volume during infection, infection duration, inducer concentration and timing of inducer addition were studied in monolayer culture. These results were then used to establish conditions for studies in microcarrier spinner flask and 1.5 L bioreactor cultures. Production processes utilizing the selected conditions will be described, together with an evaluation of the expression system.
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Expression of EGFP Reporter Protein with a Recombinant Vaccinia Virus - Comparison of Microcarrier and Cell Suspension Based Bioreactor Systems
Nicole A. Bleckwenna,b,c, William Bentleyb,c, and Joseph Shiloacha
USA, 2003
a Biotechnology Unit, NIDDK, National Institutes of Health, DHHS, Bethesda, MD
b Center for Biosystems Research, UMBI, College Park, MD
c Department of Chemical Engineering, UMCP, College Park, MD
Abstract: A recombinant vaccinia virus expression system was studied as an alternative method to produce recombinant proteins in a scaleable bioreactor system. A recombinant vaccinia virus containing the gene for the reporter protein, enhanced green fluorescent protein (EGFP), was used to study the system parameters that affect final protein expression. Evaluations of a microcarrier based method for adherent cell growth and infection in a bioreactor were performed. Controllable parameters such as temperature and dissolved oxygen level during production were tested to observe the effect on the level of protein production. Additionally, a cell suspension system was compared to the microcarrier method to determine if the levels of cell growth, viral infection, and ultimately protein expression could be maintained with a simpler production scheme.
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PUBLISHED ARTICLES ON THE ATF SYSTEM
Introduction to the ATF System and it's advantages (in German)
Dr. Christian Hetzel, P&A-Kompendium November 2009
This article highlights some of the important technologies for the future.
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CMC ICOS Biologics, USA 2009
Jason Carstens, Director Upstream Process Development, Seattle, USA
CMC Biopharmaceuticals, DK 2009
Provides an introduction to perfusion as mode of manufacturing for biotherapeutics including the challenges and advantages, plus this shows an overview of the economics. This presentation was originally shown as a webcast via the Bioprocess International webinar series.
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Use of Perfusion Technology on the Rise – New Modes Are Beginning to Gain Ground on Fed-batch Strategy
Bruce Jon Compton, Ph.D., Jacob Peter Jensen
CMC Biopharmaceuticals, DK 2007
CMC Biopharmaceuticals (owner of CMC ICOS Biologics) is one of the front runners in the dynamic CMO industry and has pushed forwards new technology allowing its clients to benefit from lower cost of goods, and higher productivity. This article discusses how perfusion is making a comeback due to increased reliability of equipment, better and easier scalability and improved modes of operation leading to higher cell densities and higher viabilities.
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Perfusion and Fed-Batch - A Comparison of Production Methodologies
Diana Morgan & Jacob Peter Jensen
CMC Biopharmaceuticals, DK 2007
An article in this newsletter describes CMC’s adoption of the ATF System into its facility and the economic benefits of modern perfusion. A comparison of operating modes of 1000L perfusion and 20,000L fed-batch is presented.
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Genetic Engineering News Article, Bioprocessing Application Note:
Scale-up of a Cell Culture Perfusion Process: A Low-Shear Filtration System that Inhibits Filter-Membrane Fouling
Jim Furey
Refine Technology, USA 2002
The characteristics and methods of scaling up from the laboratory to manufacturing using an ATF System for perfusion are discussed, and the transmembrane pressures and filter capacity are briefly covered.
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Genetic Engineering News Article, Bioprocess Tutorial:
Continuous Cell Culture Using the ATF System: A New Way to Grow Suspension or Anchorage-Dependent Cells
Jim Furey
Refine Technology, USA 2000
An early description of the ATF System used as a cell retention device in both suspension and microcarrier-based cultures. A comparison with spin filters, internal and external, is also presented.
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APPLICATIONS
Cell Banking
Read about Cell Banking and Seed Expansion in more detail
Cell Banking / Seed Expansion Application
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Overview of Perfusion, Concentrated Perfusion and Concentrated Fed-Batch CFB™
Read about your different process options in more detail
Which Process Option is Right for Me?
A Comparison of Fed-Batch, Concentrated Fed-Batch, Perfusion and Concentrated Perfusion
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Perfusion
Read more on perfusion using the ATF System
Perfusion / Cell Retention Application
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REFINE MEDIA RESOURCES
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