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  • 1
    Electronic Resource
    Electronic Resource
    Overexpression of bcl-2, apoptosis suppressing gene: Prolonged viable culture period of hybridoma and enhanced antibody production (1995)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 48 (1995), S. 118-122 
    Details
    ISSN: 0006-3592
    Keywords: apoptosis ; bcl-2 ; hybridoma ; cell survival ; antibody productivity ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Human bcl-2 DNA was introduced into mouse hybridoma 2E3 cells and expressed at a high level by using BCMGSneo vector, which reportedly amplifies as multiple copies in the cells independently of their chromosomes. The high expression of bcl-2 in BCMGSneo-bcl-2 transfectants was confirmed by western blotting. In batch cultures, the overexpression of bcl-2 raised the maximum viable cell density by 45%, delayed the initiation of apoptosis by 2 days, and prolonged the viable culture period by 4 days. The delayed initiation of apoptosis was detected by emergence of the ladder pattern on DNA electrophoresis and increase of the dead cell number. The bcl-2 transfectants produced lgG1 fourfold per batch culture in comparison with 2E3 cells transfected with BCMGSneo but not with bcl-2: a little less than twofold due to the improved survival of the cells and more than twofold due to the enhanced lgG1 production rate per cell of the bcl-2 transfectants. The method to engineer hybridoma cells genetically with bcl-2 using BCMGSneo vector for increasing viability and productivity would be widely applied for improving antibody productivity of hybridoma cultures. © 1995 John Wiley & Sons, Inc.
    Additional Material: 5 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/bit.260480205
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  • 2
    Electronic Resource
    Electronic Resource
    Anti-apoptotic genes, bag-1 and bcl-2, enabled hybridoma cells to survive under treatment for arresting cell cycle (1997)
    Springer
    Cytotechnology 25 (1997), S. 17-23 
    Details
    ISSN: 1573-0778
    Keywords: anti-apoptotic ; bag-1 ; bcl-2 ; cell cycle arrest ; excess thymidine ; serum limitation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Hybridoma 2E3-O cells were transfected with bcl-2 alone or with bcl-2 and bag-1 in combination. The bcl-2/bag-1 transfectant survived maintaining viability above 75% for almost 5 days when the cells were treated with excess (30 mM) thymidine for arresting cell cycle, whereas the mock transfectant survived for only 2 days, and the bcl-2 alone transfectant lived for 4 days. Owing to this extended viable culture period, the bcl-2/bag-1 transfectant produced twofold amount of antibody in comparison with the mock transfectant in non-proliferating state prepared by the excess thymidine treatment. When their proliferation was arrested by serum limitation, the bcl-2/bag-1 transfectant and the bcl-2 alone transfectant survived for 3 days maintaining viability above 75% while the mock transfectant survived only 1 day. The bcl-2/bag-1 transfectans produced the antibody at the rate three times as high as the bcl-2 alone transfectant and the mock transfectant in non-proliferating state established by serum limitation. Such genetic engineering of hybridoma cells for improving survival in the non-proliferating state will be useful for using nutrients in culture medium efficiently to produce antibody, since nutrients could be diverted from cell proliferation to antibody production in such non-proliferating viable cell culture.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007954103572
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  • 3
    Electronic Resource
    Electronic Resource
    Reinforcing apoptosis-resistance of COS and myeloma cells by transfecting with bcl-2 gene (1997)
    Springer
    Cytotechnology 25 (1997), S. 25-33 
    Details
    ISSN: 1573-0778
    Keywords: apoptosis ; bcl-2 ; COS cell ; myeloma
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract COS, myeloma and HeLa cells, which are commonly used for protein production by cell culture, were transfected with human bcl-2 gene encoded on the shuttle vector BCMGS. Expression of human bcl-2 improved survival of cells remarkably, mildly, or negligibly for COS, myeloma, and HeLa, respectively. Four clones were obtained from the human bcl-2 expressing cell population of COS cells. They expressed human bcl-2 almost at the same level. The viable cell numbers were 6, 2.5, 2.5, and 0.8 times as many for the clones #8, #5, #6, and #7, respectively, as for the control COS cells, when they were cultured at low (0.2%) serum concentration for 9 days. The bcl-2 overexpressing COS cells showed morphology different from that of the control COS cells in serum limited condition. When transfected with mouse lambda protein gene carried by an SV40-derived vector, clone #8 of the bcl-2 transfected COS cells continued the transient expression of lambda protein longer than the control COS cells.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007935026770
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  • 4
    Electronic Resource
    Electronic Resource
    Characterization and fed-batch culture of hybridoma overexpressing apoptosis suppressing gene bcl-2 (1997)
    Springer
    Cytotechnology 24 (1997), S. 135-141 
    Details
    ISSN: 1573-0778
    Keywords: antibody productivity ; apoptosis ; bcl-2 ; fed batchculture ; hybridoma
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Mouse hybridoma 2E3 transfected with human bcl-2 gene survived longer with increasing expression level of bcl-2 when cultured in DME medium supplemented with 9% serum. One of the transfectants, 2E3BCMGbcl-2, overexpressed bcl-2 and could maintain viable cell density higher than the initial density for more than four days at a low 0.5% serum concentration. In comparison a mock transfectant 2E3BCMG remained viable for only one day. However, both hybridomas died out within a day in serum-free medium. These results suggested that bcl-2 needed a small amount of some serum components to suppress apoptosis of the hybridoma. Overexpression of bcl-2 also suppressed apoptosis of the hybridoma induced by glutamine deprivation. When hybridoma 2E3BCMGbcl-2 was inoculated in DME medium supplemented with 9% serum and cultured for 10 d with additional 2% serum feed at day 4 of the culture, viable cell density increased 2-fold and antibody produced 3-fold, in comparison with mock transfected 2E3 cultured in the same manner. The mock transfectant with additional feed of serum at day 4 of the culture showed no difference in viable cell density and antibody production. These results suggested that the mock transfectant committed to apoptosis before day 4 of the culture and the additional serum at day 4 could not reverse the commitment.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007922104344
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