Process development of human multipotent stromal cell microcarrier culture using an automated high-throughput microbioreactor

Qasim A. Rafiq, Mariana P. Hanga, Thomas R.J. Heathman, Karen Coopman, Alvin W. Nienow, David J. Williams, Christopher J. Hewitt

Research output: Contribution to journalArticle

Abstract

Microbioreactors play a critical role in process development as they reduce reagent requirements and can facilitate high-throughput screening of process parameters and culture conditions. Here we have demonstrated and explained in detail, for the first time, the amenability of the automated ambr15 cell culture microbioreactor system for the development of scalable adherent human mesenchymal multipotent stromal/stem cell (hMSC) microcarrier culture processes. This was achieved by first improving suspension and mixing of the microcarriers and then improving cell attachment thereby reducing the initial growth lag phase. The latter was achieved by using only 50% of the final working volume of medium for the first 24 h and using an intermittent agitation strategy. These changes resulted in > 150 % increase in viable cell density after 24 h compared to the original process (no agitation for 24 h and 100 % working volume). Using the same methodology as in the ambr15, similar improvements were obtained with larger scale spinner flask studies. Finally, this improved bioprocess methodology based on a serum-based medium was applied to a serum-free process in the ambr15, resulting in > 250% increase in yield compared to the serum-based process. At both scales, the agitation used during culture was the minimum required for microcarrier suspension, NJS. The use of the ambr15, with its improved control compared to the spinner flask, reduced the coefficient of variation on viable cell density in the serum containing medium from 7.65% to 4.08%, and the switch to serum free further reduced these to 1.06% and 0.54% respectively. The combination of both serum-free and automated processing improved the reproducibility more than 10-fold compared to the serum-based, manual spinner flask process. The findings of this study demonstrate that the ambr15 microbioreactor is an effective tool for bioprocess development of hMSC microcarrier cultures and that a combination of serum-free medium, control and automation improves both process yield and consistency.
Original languageEnglish
Pages (from-to)2253-2266
JournalBiotechnology and Bioengineering
Volume114
Issue number10
Early online date19 Jun 2017
DOIs
Publication statusPublished - 1 Oct 2017

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Human Development
Stromal Cells
Cell culture
Cell Culture Techniques
Throughput
Stem cells
Suspensions
Serum
Multipotent Stem Cells
Serum-Free Culture Media
Mesenchymal Stromal Cells
Screening
Automation
Switches
Cell Count
Processing
Growth

Bibliographical note

Funding: EPSRC (EP/I017801/1)

Keywords

  • human mesenchymal multipotent stromal cell
  • bioprocessing
  • microcarrier
  • microbioreactor
  • cell therapy
  • scale down

Cite this

Rafiq, Qasim A. ; Hanga, Mariana P. ; Heathman, Thomas R.J. ; Coopman, Karen ; Nienow, Alvin W. ; Williams, David J. ; Hewitt, Christopher J. / Process development of human multipotent stromal cell microcarrier culture using an automated high-throughput microbioreactor. In: Biotechnology and Bioengineering . 2017 ; Vol. 114, No. 10. pp. 2253-2266.
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Process development of human multipotent stromal cell microcarrier culture using an automated high-throughput microbioreactor. / Rafiq, Qasim A.; Hanga, Mariana P.; Heathman, Thomas R.J.; Coopman, Karen; Nienow, Alvin W.; Williams, David J.; Hewitt, Christopher J.

In: Biotechnology and Bioengineering , Vol. 114, No. 10, 01.10.2017, p. 2253-2266.

Research output: Contribution to journalArticle

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T1 - Process development of human multipotent stromal cell microcarrier culture using an automated high-throughput microbioreactor

AU - Rafiq, Qasim A.

AU - Hanga, Mariana P.

AU - Heathman, Thomas R.J.

AU - Coopman, Karen

AU - Nienow, Alvin W.

AU - Williams, David J.

AU - Hewitt, Christopher J.

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Y1 - 2017/10/1

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AB - Microbioreactors play a critical role in process development as they reduce reagent requirements and can facilitate high-throughput screening of process parameters and culture conditions. Here we have demonstrated and explained in detail, for the first time, the amenability of the automated ambr15 cell culture microbioreactor system for the development of scalable adherent human mesenchymal multipotent stromal/stem cell (hMSC) microcarrier culture processes. This was achieved by first improving suspension and mixing of the microcarriers and then improving cell attachment thereby reducing the initial growth lag phase. The latter was achieved by using only 50% of the final working volume of medium for the first 24 h and using an intermittent agitation strategy. These changes resulted in > 150 % increase in viable cell density after 24 h compared to the original process (no agitation for 24 h and 100 % working volume). Using the same methodology as in the ambr15, similar improvements were obtained with larger scale spinner flask studies. Finally, this improved bioprocess methodology based on a serum-based medium was applied to a serum-free process in the ambr15, resulting in > 250% increase in yield compared to the serum-based process. At both scales, the agitation used during culture was the minimum required for microcarrier suspension, NJS. The use of the ambr15, with its improved control compared to the spinner flask, reduced the coefficient of variation on viable cell density in the serum containing medium from 7.65% to 4.08%, and the switch to serum free further reduced these to 1.06% and 0.54% respectively. The combination of both serum-free and automated processing improved the reproducibility more than 10-fold compared to the serum-based, manual spinner flask process. The findings of this study demonstrate that the ambr15 microbioreactor is an effective tool for bioprocess development of hMSC microcarrier cultures and that a combination of serum-free medium, control and automation improves both process yield and consistency.

KW - human mesenchymal multipotent stromal cell

KW - bioprocessing

KW - microcarrier

KW - microbioreactor

KW - cell therapy

KW - scale down

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