Document Detail


CHO-K1 host cells adapted to growth in glutamine-free medium by FACS-assisted evolution.
MedLine Citation:
PMID:  20931603     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
During the process of recombinant cell line optimisation for production of biopharmaceuticals, multiple cellular properties like robustness against stress, the attainment of high cell concentrations and maintenance of high viability must be considered to maximize protein yield. To improve growth and viability, glutamine is supplemented as an alternative energy source for rapidly dividing cells that oxidize glucose inefficiently. However, the resulting by-product ammonia is toxic at high concentrations and has a negative impact on protein glycosylation, a major quality-determining parameter of biopharmaceuticals. In this work, the CHO-K1 cell line was adapted to a chemically defined medium and suspension growth within 3 weeks. Subsequently, the glutamine concentration was stepwise reduced from 8 to 4 and 2 mM. After each reduction, both the final cell concentration in the batch and the viability decreased. To force a rapid evolution of cells to achieve high final cell concentrations, cells were seeded at high densities (10(7) cells/mL) and surviving cells were sorted by FACS or MACS when viability declined to 10% (typically after 24 h). Sorted cells were grown in batch until viability declined to 10% and viable cells recovered again. The final sorted population was able to reach comparable or even better viable cell concentrations and showed a significantly improved viability compared to their ancestors. The 2 mM glutamine-adapted cell line was directly transferred into glutamine-free medium and was able to grow at comparable rates without requiring further adaptation. Cells compensated the lack of glutamine by increasing their consumption of glutamate and aspartate.
Authors:
Juan A Hernández Bort; Beate Stern; Nicole Borth
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Publication Detail:
Type:  Journal Article    
Journal Detail:
Title:  Biotechnology journal     Volume:  5     ISSN:  1860-7314     ISO Abbreviation:  Biotechnol J     Publication Date:  2010 Oct 
Date Detail:
Created Date:  2010-10-08     Completed Date:  2011-01-24     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  101265833     Medline TA:  Biotechnol J     Country:  Germany    
Other Details:
Languages:  eng     Pagination:  1090-7     Citation Subset:  IM    
Affiliation:
Department of Biotechnology, University of Natural Resources and Applied Life Sciences, Vienna, Austria.
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MeSH Terms
Descriptor/Qualifier:
Animals
CHO Cells / cytology*,  drug effects*
Cell Survival / drug effects
Cricetinae
Cricetulus
Culture Media / chemistry,  pharmacology*
Flow Cytometry
Glutamine / chemistry*
Chemical
Reg. No./Substance:
0/Culture Media; 56-85-9/Glutamine

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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