Document Detail

Stability of producer hybridoma cell lines after cell sorting: a case study.
MedLine Citation:
PMID:  7764937     Owner:  NLM     Status:  MEDLINE    
Flow cytometry was used in combination with immunofluorescent staining for intracellular and surface-associated antibody contents to identify a significant non-producer cell fraction in a murine hybridoma cell line that had shown a decline in monoclonal antibody productivity with passaging. Viable producer cells stained for surface-associated antibody content were isolated by cell sorting on the basis of surface fluorescence intensity. The fraction of nonproducers was initially reduced from 85% to 20%. Sorting a second time, after these cells were cultivated for 2 weeks, further reduced the fraction of nonproducers to less than 4%. The stability of this purified producer hybridoma cell line during passaging and after a freeze-thaw cycle was investigated. This cell line was found to be highly unstable. A simple batch-growth model simulation was used as follows: (i) to demonstrate that nonproducers appear in hybridoma cell lines after a rare, random mutation event that results in the loss of the heavy-chain gene and/or light-chain gene expression; (ii) to show that a high rate of conversion of producer hybridomas to nonproducer hybridomas cannot entirely explain the population dynamics; (iii) to estimate the rate of conversion of producers to nonproducers to be 8.7 x 10(-5) h-1; (iv) to show that, for this conversion rate, the nonproducer cells' specific growth rates need only be 9% higher than those of the producer cells to dominate the hybridoma culture after only 25 passages; and (v) to predict that producer cells are preferentially lost during cell freezing and thawing procedures. The data suggest that particularly unstable cell lines should be analyzed and purified frequently to prevent overgrowth by nonproducers.
S J Kromenaker; F Srienc
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, Non-P.H.S.    
Journal Detail:
Title:  Biotechnology progress     Volume:  10     ISSN:  8756-7938     ISO Abbreviation:  Biotechnol. Prog.     Publication Date:    1994 May-Jun
Date Detail:
Created Date:  1994-08-03     Completed Date:  1994-08-03     Revised Date:  2006-11-15    
Medline Journal Info:
Nlm Unique ID:  8506292     Medline TA:  Biotechnol Prog     Country:  UNITED STATES    
Other Details:
Languages:  eng     Pagination:  299-307     Citation Subset:  B    
Department of Chemical Engineering and Materials Science, University of Minnesota, Minneapolis 55455.
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MeSH Terms
Antibodies, Monoclonal / biosynthesis
Cell Cycle / physiology
Cell Division / physiology
Cell Separation
Flow Cytometry
Fluorescent Antibody Technique
Hybridomas / cytology*,  immunology*
Reg. No./Substance:
0/Antibodies, Monoclonal

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