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Publications (9 of 9) Show all publications
Al-Khalili, L., Gillner, K., Zhang, Y., Åstrand, C., Shokri, A., Hughes-Brittain, N., . . . Chotteau, V. (2016). Characterization of Human CD133+Cells in Biocompatible Poly(l-lactic acid) Electrospun Nano-Fiber Scaffolds. Journal of Biomaterials and Tissue Engineering, 6(12), 959-966
Open this publication in new window or tab >>Characterization of Human CD133+Cells in Biocompatible Poly(l-lactic acid) Electrospun Nano-Fiber Scaffolds
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2016 (English)In: Journal of Biomaterials and Tissue Engineering, ISSN 2157-9083, E-ISSN 2157-9091, Vol. 6, no 12, p. 959-966Article in journal (Refereed) Published
Abstract [en]

CD133+ cells are potential myogenic progenitors for skeletal muscle regeneration to treat muscular dystrophies. The proliferation of human CD133+ stem cells was studied for 14 days in 3D biomimetic electrospun poly-L-lactic acid (PLLA) nano-fiber scaffolds. Additionally, the myogenic differentiation of the cells was studied during the last 7 days of the culture period. The cells were homogeneously distributed in the 3D scaffolds while colony formation and myotube formation occurred in 2D. After a lag phase due to lower initial cell attachment and an adaptation period, the cell growth rate in 3D was comparable to 2D after 7 and 14 days of culture. The expression of the stem cell (SC) marker PAX7 was 1.5-fold higher in 3D than 2D while the differentiation markers MyoG, Desmin and MyoD were only slightly changed (or remain unchanged) in 3D but strongly increased in 2D (12.6, 3.9, and 7.9-fold), and the myotube formation observed in 2D was absent in 3D. The marker expression during proliferation and differentiation, together with the absence of myotubes in 3D, indicates a better maintenance of stemness in 3D PLLA and stronger tendency for spontaneous differentiation in 2D culture. This makes 3D PLLA a promising biomaterial for the expansion of functional CD133+ cells.

Place, publisher, year, edition, pages
American Scientific Publishers, 2016
Keywords
Myogenic Progenitor Cell, CD133+Cells, Myogenic Differentiation, 3D Cell Culturing, Electrospun Biodegradable Nano-Fiber Scaffold
National Category
Biological Sciences
Identifiers
urn:nbn:se:kth:diva-198952 (URN)10.1166/jbt.2016.1531 (DOI)000387148500005 ()2-s2.0-84998636398 (Scopus ID)
Funder
EU, FP7, Seventh Framework Programme
Note

QC 20170113

Available from: 2017-01-13 Created: 2016-12-22 Last updated: 2017-11-29Bibliographically approved
Zhang, Y. & Chotteau, V. (2015). Observation of Chinese Hamster Ovary Cells retained inside the non-woven fiber matrix of the CellTank bioreactor. Data in Brief, 5, 586-588
Open this publication in new window or tab >>Observation of Chinese Hamster Ovary Cells retained inside the non-woven fiber matrix of the CellTank bioreactor
2015 (English)In: Data in Brief, ISSN 2352-3409, Vol. 5, p. 586-588Article in journal (Refereed) Published
Abstract [en]

This data article shows how the recombinant Chinese Hamster Ovary (CHO) cells are located in the interstices of the matrix fibers of a CellTank bioreactor after completion of a perfusion culture, supporting the article entitled "Very high cell density perfusion of CHO cells anchored in a non-woven matrix-based bioreactor" by Zhang et al. [1]. It provides a visualization of the cell distribution in the non-woven fiber matrix in a deeper view.

Place, publisher, year, edition, pages
Elsevier, 2015
National Category
Cell and Molecular Biology
Identifiers
urn:nbn:se:kth:diva-181839 (URN)10.1016/j.dib.2015.10.006 (DOI)2-s2.0-84945243514 (Scopus ID)
Note

QC 20160211

Available from: 2016-02-11 Created: 2016-02-05 Last updated: 2018-01-10Bibliographically approved
Zhang, Y., Stobbe, P., Silvander, C. O. & Chotteau, V. (2015). Very high cell density perfusion of CHO cells anchored in a non-woven matrix-based bioreactor. Journal of Biotechnology, 213, 28-41
Open this publication in new window or tab >>Very high cell density perfusion of CHO cells anchored in a non-woven matrix-based bioreactor
2015 (English)In: Journal of Biotechnology, ISSN 0168-1656, E-ISSN 1873-4863, Vol. 213, p. 28-41Article in journal (Refereed) Published
Abstract [en]

Recombinant Chinese Hamster Ovary (CHO) cells producing IgG monoclonal antibody were cultivated in a novel perfusion culture system CellTank, integrating the bioreactor and the cell retention function. In this system, the cells were harbored in a non-woven polyester matrix perfused by the culture medium and immersed in a reservoir. Although adapted to suspension, the CHO cells stayed entrapped in the matrix. The cell-free medium was efficiently circulated from the reservoir into- and through the matrix by a centrifugal pump placed at the bottom of the bioreactor resulting in highly homogenous concentrations of the nutrients and metabolites in the whole system as confirmed by measurements from different sampling locations. A real-time biomass sensor using the dielectric properties of living cells was used to measure the cell density. The performances of the CellTank were studied in three perfusion runs. A very high cell density measured as 200 pF/cm (where 1 pF/cm is equivalent to 1 x 106 viable cells/mL) was achieved at a perfusion rate of 10 reactor volumes per day (RV/day) in the first run. In the second run, the effect of cell growth arrest by hypothermia at temperatures lowered gradually from 37 C to 29 C was studied during 13 days at cell densities above 100 pF/cm. Finally a production run was performed at high cell densities, where a temperature shift to 31 C was applied at cell density 100 pF/cm during a production period of 14 days in minimized feeding conditions. The IgG concentrations were comparable in the matrix and in the harvest line in all the runs, indicating no retention of the product of interest. The cell specific productivity was comparable or higher than in Erlenmeyer flask batch culture. During the production run, the final harvested IgG production was 35 times higher in the CellTank compared to a repeated batch culture in the same vessel volume during the same time period.

Place, publisher, year, edition, pages
[Zhang, Ye; Chotteau, Veronique] KTH, Royal Inst Technol, Cell Technol Grp CETEG, Sch Biotechnol,Dept Ind Biotechnol Bioproc Design, SE-10691 Stockholm, Sweden. [Stobbe, Per] PerfuseCell, DK-2840 Holte, Denmark. [Silvander, Christian Orrego] Belach Biotekn, SE-14250 Skogas, Sweden.: , 2015
Keywords
Disposable bioreactor, On-line biomass sensor, IgG production, Dielectric spectroscopy, Hypothermia
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
urn:nbn:se:kth:diva-176326 (URN)10.1016/j.jbiotec.2015.07.006 (DOI)000362286400005 ()26211737 (PubMedID)2-s2.0-84942837005 (Scopus ID)
Note

QC 20151117

Available from: 2015-11-17 Created: 2015-11-03 Last updated: 2017-05-23Bibliographically approved
Chotteau, V., Zhang, Y. & Clincke, M.-F. (2014). Very High Cell Density in Perfusion of CHO Cells by ATF, TFF, Wave Bioreactor, and/or CellTank Technologies: Impact of Cell Density and Applications. In: Ganapathy Subramanian (Ed.), Continuous Processing in Pharmaceutical Manufacturing: (pp. 339-356). Germany Weinheim: Wiley-VCH Verlagsgesellschaft
Open this publication in new window or tab >>Very High Cell Density in Perfusion of CHO Cells by ATF, TFF, Wave Bioreactor, and/or CellTank Technologies: Impact of Cell Density and Applications
2014 (English)In: Continuous Processing in Pharmaceutical Manufacturing / [ed] Ganapathy Subramanian, Germany Weinheim: Wiley-VCH Verlagsgesellschaft, 2014, p. 339-356Chapter in book (Other academic)
Place, publisher, year, edition, pages
Germany Weinheim: Wiley-VCH Verlagsgesellschaft, 2014
Keywords
CellTank, perfusion, filtration, ultra filtration, Chinese hamster ovary (CHO) cell, high-cell density perfusion processes, monoclonal antibody, ATF, TFF
National Category
Bioprocess Technology
Identifiers
urn:nbn:se:kth:diva-166113 (URN)10.1002/9783527673681.ch13 (DOI)2-s2.0-84949057950 (Scopus ID)9783527673681 (ISBN)
Note

QC 20150508

Available from: 2015-05-01 Created: 2015-05-01 Last updated: 2015-05-08Bibliographically approved
Clincke, M.-F., Mölleryd, C., Zhang, Y., Lindskog, E., Walsh, K. & Chotteau, V. (2013). Very high density of CHO cells in perfusion by ATF or TFF in WAVE bioreactor. Part I. Effect of the cell density on the process. Biotechnology progress (Print), 29(3), 754-767
Open this publication in new window or tab >>Very high density of CHO cells in perfusion by ATF or TFF in WAVE bioreactor. Part I. Effect of the cell density on the process
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2013 (English)In: Biotechnology progress (Print), ISSN 8756-7938, E-ISSN 1520-6033, Vol. 29, no 3, p. 754-767Article in journal (Refereed) Published
Abstract [en]

High cell density perfusion process of antibody producing CHO cells was developed in disposable WAVE Bioreactor using external hollow fiber filter as cell separation device. Both classical tangential flow filtration (TFF) and alternating tangential flow system (ATF) equipment were used and compared. Consistency of both TFF- and ATF-based cultures was shown at 20-35 x 106 cells/mL density stabilized by cell bleeds. To minimize the nutrients deprivation and by-product accumulation, a perfusion rate correlated to the cell density was applied. The cells were maintained by cell bleeds at density 0.9-1.3 x 108 cells/mL in growing state and at high viability for more than 2 weeks. Finally, with the present settings, maximal cell densities of 2.14 x 108 cells/mL, achieved for the first time in a wave-induced bioreactor, and 1.32 x 108 cells/mL were reached using TFF and ATF systems, respectively. Using TFF, the cell density was limited by the membrane capacity for the encountered high viscosity and by the pCO2 level. Using ATF, the cell density was limited by the vacuum capacity failing to pull the highly viscous fluid. Thus, the TFF system allowed reaching higher cell densities. The TFF inlet pressure was highly correlated to the viscosity leading to the development of a model of this pressure, which is a useful tool for hollow fiber design of TFF and ATF. At very high cell density, the viscosity introduced physical limitations. This led us to recommend cell densities under 1.46 x 108 cell/mL based on the analysis of the theoretical distance between the cells for the present cell line.

Place, publisher, year, edition, pages
Wiley-Blackwell, 2013
Keywords
wave bioreactor, alternating tangential flow hollow fiber, tangential flow filtration hollow fiber, perfusion, cell broth viscosity, Chinese Hamster Ovary cell
National Category
Biological Sciences
Identifiers
urn:nbn:se:kth:diva-124990 (URN)10.1002/btpr.1704 (DOI)000320387300019 ()2-s2.0-84879258603 (Scopus ID)
Note

QC 20130806

Available from: 2013-08-06 Created: 2013-08-02 Last updated: 2017-05-23Bibliographically approved
Clincke, M.-F., Mölleryd, C., Samani, P. K., Zhang, Y., Lindskog, E., Walsh, K. & Chotteau, V. (2011). Perfusion of an IgG producing CHO cell line at very high cell density by ATF or by TFF in WAVE Bioreactor™. Paper presented at Bioprocessing Summit, Boston, MA, USA, Aug. 22-25, 2011 - Optimizing Cell Culture Technology.
Open this publication in new window or tab >>Perfusion of an IgG producing CHO cell line at very high cell density by ATF or by TFF in WAVE Bioreactor™
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2011 (English)Conference paper, Published paper (Other academic)
Abstract [en]

Perfusion of an IgG producing CHO cell line was performed in a WAVE Bioreactor™ using either Alternating Tangential Flow or Tangential Flow Filtration. The properties and performances obtained with both filtration systems were compared. Very high cell densities were achieved and could be stably maintained. Then the cell density could be significantly further increased showing the capacity of the system set-up.

Keywords
perfusion, hollow fiber filter, perfusion, ATF, TFF, CHO cells
National Category
Medical Biotechnology
Research subject
SRA - Molecular Bioscience
Identifiers
urn:nbn:se:kth:diva-87428 (URN)
Conference
Bioprocessing Summit, Boston, MA, USA, Aug. 22-25, 2011 - Optimizing Cell Culture Technology
Note
Invited speaker at the Bioprocessing Summit, Boston, MA, USA, Aug. 22-25, 2011 - Optimizing Cell Culture Technology. QC 20120319Available from: 2012-02-14 Created: 2012-02-14 Last updated: 2012-03-19Bibliographically approved
Clincke, M.-F., Mölleryd, C., Samani, P. K., Zhang, Y., Lindskog, E., Walsh, K. & Chotteau, V. (2011). Perfusion of an IgG producing CHO cell line at very high cell density by ATF or by TFF in WAVE Bioreactor™. Paper presented at HansonWade Conference Optimizing Cell Line Performance, July, 26-28, 2011, London, UK.
Open this publication in new window or tab >>Perfusion of an IgG producing CHO cell line at very high cell density by ATF or by TFF in WAVE Bioreactor™
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2011 (English)Conference paper, Published paper (Other academic)
Abstract [en]

Perfusion of an IgG producing CHO cell line was performed in a WAVE Bioreactor™ using either Alternating Tangential Flow or Tangential Flow Filtration. The properties and performances obtained in this bioreactor with both filtration systems were studied.

  • Very high cell densities were achieved and could be stably maintained at high viability indicating of a healthy process suitable for instance for efficient cell banking.

Cell density could be significantly further increased showing the capacity of the system set-up.

Keywords
perfusion, hollow fiber filter, perfusion, ATF, TFF, CHO cells
National Category
Medical Biotechnology
Research subject
SRA - Molecular Bioscience
Identifiers
urn:nbn:se:kth:diva-87442 (URN)
Conference
HansonWade Conference Optimizing Cell Line Performance, July, 26-28, 2011, London, UK
Note
Invited speaker at the Hanson Wade Conference Optimizing Cell Line Performance, July, 26-28, 2011, London, UK. QC 20130319Available from: 2012-02-14 Created: 2012-02-14 Last updated: 2012-03-19Bibliographically approved
Clincke, M.-F., Mölleryd, C., Zhang, Y., Lindskog, E., Walsh, K. & Chotteau, V. (2011). Study of a recombinant CHO cell line producing a monoclonal antibody by ATF or TFF external filter perfusion in a WAVE Bioreactor™. In: BMC Proceedings, 2011, Volume 5, Supplement 8, P105. Paper presented at 22d ESACT (European Society for Animal Cell Technology), May 15-18, 2011, Vienna, Austria (pp. 105). BioMed Central
Open this publication in new window or tab >>Study of a recombinant CHO cell line producing a monoclonal antibody by ATF or TFF external filter perfusion in a WAVE Bioreactor™
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2011 (English)In: BMC Proceedings, 2011, Volume 5, Supplement 8, P105, BioMed Central, 2011, p. 105-Conference paper, Published paper (Refereed)
Abstract [en]

Major advantages of perfusion are high cell numbers and high total production in a relatively small size bioreactor. Moreover, perfusion is optimal when the product of interest is unstable or if the product yield is low. On the other hand, disadvantages are for example technical challenges originating from non-robust cell separation devices as well as sterility concerns from the more complex set-up needed.

In the present work, the use of a WAVE Bioreactor™ system 20/50 in perfusion mode with10 L disposable Cellbag™ bioreactors customized with two dip tubes in combination with disposable hollow fiber filters as external cell separating devices were investigated. A comparison between Alternating Tangential Flow (ATF) and Tangential Flow Filtration (TFF) was performed using a recombinant CHO cell line producing a monoclonal antibody (mAb) as a model system. 

Place, publisher, year, edition, pages
BioMed Central, 2011
Keywords
perfusion, ATF, TFF, hollow fiber, cell separation, Chinese Hamster Ovary, wave bioreactor, high cell density
National Category
Pharmaceutical Biotechnology
Research subject
SRA - Molecular Bioscience
Identifiers
urn:nbn:se:kth:diva-86387 (URN)10.1186/1753-6561-5-S8-P105 (DOI)
Conference
22d ESACT (European Society for Animal Cell Technology), May 15-18, 2011, Vienna, Austria
Note
QC 20120214Available from: 2012-02-14 Created: 2012-02-13 Last updated: 2012-03-22Bibliographically approved
Schwarz, H., Zhang, Y., Zhan, C., Malm, M., Field, R., Turner, R., . . . Chotteau, V.Small-scale bioreactor supports high density HEK293 cell perfusion culture for the production of recombinant Erythropoietin.
Open this publication in new window or tab >>Small-scale bioreactor supports high density HEK293 cell perfusion culture for the production of recombinant Erythropoietin
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(English)Manuscript (preprint) (Other academic)
Abstract [en]

Process intensification in mammalian cell culture-based recombinant protein production has been achieved by high cell density perfusion exceeding 108 cells/mL in the recent years. As the majority of therapeutic proteins are produced in Chinese Hamster Ovary (CHO) cells, intensified perfusion processes have been mainly developed for this type of host cell line. However, the use of CHO cells can result in non-human posttranslational modifications of the protein of interest, which may be disadvantageous compared with human cell lines.

In this study, we developed a high cell density perfusion process of Human Embryonic Kidney (HEK293) cells producing recombinant human Erythropoietin (rhEPO). Firstly, a small-scale perfusion system from commercial bench-top screening bioreactors was developed for <250 mL working volume. Then, after the first trial runs with CHO cells, the system was modified for HEK293 cells (more sensitive than CHO cells) to achieve a higher oxygen transfer under mild aeration and agitation conditions. Steady states for medium (20 x 106 cells/mL) and high cell densities (80 x 106 cells/mL), normal process temperature (37 °C) and mild hypothermia (33 °C) as well as different cell specific perfusion rates (CSPR) from 10 to 60 pL/cell/day were applied to study the performance of the culture. The volumetric productivity was maximized for the high cell density steady state but decreased when an extremely low CSPR of 10 pL/cell/day was applied. The shift from high to low CSPR strongly reduced the nutrient uptake rates. The results from our study show that human cell lines, such as HEK293 can be used for intensified perfusion processes. 

Keywords
Chinese Hamster Ovary (CHO) cells, Erythropoietin, High cell density culture, Human Embryonic Kidney 239 (HEK293) cells, Perfusion process, Small-scale bioreactor
National Category
Industrial Biotechnology
Identifiers
urn:nbn:se:kth:diva-252022 (URN)
Note

QC 20190523

Available from: 2019-05-23 Created: 2019-05-23 Last updated: 2019-05-23Bibliographically approved
Organisations
Identifiers
ORCID iD: ORCID iD iconorcid.org/0000-0002-0841-8845

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