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  • 1
    Electronic Resource
    Electronic Resource
    Measurement of hybridoma cell number, viability, and morphology using fully automated image analysis
    Amsterdam : Elsevier
    Enzyme and Microbial Technology 16 (1994), S. 29-35 
    Details
    ISSN: 0141-0229
    Keywords: Hybridoma ; cell morphology ; cell viability ; image analysis
    Source: Elsevier Journal Backfiles on ScienceDirect 1907 - 2002
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1016/0141-0229(94)90106-6
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  • 2
    Electronic Resource
    Electronic Resource
    Hybridoma growth and monoclonal antibody production in a dialysis perfusion system
    Amsterdam : Elsevier
    Enzyme and Microbial Technology 16 (1994), S. 688-695 
    Details
    ISSN: 0141-0229
    Keywords: Hybridomas ; dialysis ; mathematical model ; metabolism ; monoclonal antibody ; perfusion
    Source: Elsevier Journal Backfiles on ScienceDirect 1907 - 2002
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1016/0141-0229(94)90091-4
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  • 3
    Electronic Resource
    Electronic Resource
    Quantitative cytochemical analysis of immobilised hybridoma cells (1989)
    Springer
    Applied microbiology and biotechnology 31 (1989), S. 430-433 
    Details
    ISSN: 1432-0614
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Summary The activity of hybridoma cells immobilised in agarose gels is observed to decline with time in a manner consistent with oxygen diffusion limitation. The final thickness of the active cell layer is dependent on the initial cell density. The effect of diffusion resistance is more pronounced in medium without serum.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00257617
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  • 4
    Electronic Resource
    Electronic Resource
    Estimation of disruption of animal cells by laminar shear stress (1992)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 40 (1992), S. 1004-1010 
    Details
    ISSN: 0006-3592
    Keywords: mammalian cell ; disruption ; shear stress ; mechanical properties ; micromanipulation ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Using mechanical cell properties measured by micromanipulation, and a model of cell distortion in laminar flow fields, a method has been developed for predicting disruption of animal cells by laminar shear stresses. Predictions of the model were compared with measured losses of cell number and viability of TB/C3 murine hybridomas sheared in a cone and plate viscometer at shear rates up to 3950 s-1, and shear stresses up to 600 Nm-2, achieved by enhancement of viscosity with dextran. In all cases, the experimental, results and predictions were within 30%. Such excellent agreement suggests it might be possible to use micromanipulation measurements of animal cell mechanical properties to predict cell damage in more complex flow fields, such as those in bioreactors. © 1992 John Wiley & Sons, Inc.
    Additional Material: 7 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/bit.260400903
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  • 5
    Electronic Resource
    Electronic Resource
    Cell death in bioreactors: A role for apoptosis (1994)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 44 (1994), S. 720-726 
    Details
    ISSN: 0006-3592
    Keywords: cell death ; apoptosis ; hybridoma cells ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The incidence of apoptotic and necrotic cell death was compared in CHO, SF9 insect cells and murine plasmacytoma (J558L) and hybridoma (TB/C3) cells during in vitro cultivation in batch cultures. Acridine orange staining and fluorescence microscopy enabled the visualization of a classic morphological feature of apoptotic cell, the presence of condensed and/or fragmented chromatin. DNA gel electrophoresis was employed to show an additional characteristic of the process, the endonuclease-mediated fragmentation of DNA into multiples of 180 base pairs. The levels of apoptosis at the end of batch cultures of plasmacytoma and hybridoma cell lines were found to be 60% and 90% of total dead cells, respectively. However, employing the above-mentioned techniques, the biochemical and morphological features of apoptosis were not found in CHO and SF9 insect cells. Some factors affecting the induction of apoptosis during the batch culture of the hybridoma and plasmacytoma cell lines were identified. The most effective inducer was found to be glutamine limitation, followed by (in order of importance) serum limitation, glucose limitation, and ammonia toxicity. Blockage of the cell cycle of the plasmacytoma and hybridoma cells using thymidine resulted in the induction of apoptosis. This has important implications for the development of cell culture processes that minimize cell division and thereby increase specific productivity. © 1994 John Wiley & Sons, Inc.
    Additional Material: 7 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/bit.260440608
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  • 6
    Electronic Resource
    Electronic Resource
    Estimation of disruption of animal cells by turbulent capillary flow (1993)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 42 (1993), S. 987-993 
    Details
    ISSN: 0006-3592
    Keywords: cell disruption ; turbulent flow ; mammalian cells ; mechanical properties ; micromanipulation ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Disruption of animal cells in turbulent capillary flows has been predicted from a model of cell-hydrodynamic interactions using cell mechanical properties determined by micromanipulation. Eddies of sizes similar to or smaller than the cells are presumed to interact with those cells, causing local surface deformations. The proposed mechanism of cell damage is that such deformations result in an increase in membrane tension and surface energy and that a cell disrupts when its bursting membrane tension and bursting surface energy are exceeded. The surface energy of the cells is estimated from the kinetic energy of appropriately sized eddies. To test the model, cells were disrupted in turbulent flows in capillaries at mean energy dissipation rates up to 2 × 104 m2/s3. In all cases the model underestimated the cell disruption by about 15%. Such good agreement implies that the approach of the model to the complicated phenomena of cell turbulence interactions is reasonable. © 1993 John Wiley & Sons, Inc.
    Additional Material: 2 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/bit.260420809
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  • 7
    Electronic Resource
    Electronic Resource
    Death mechanisms of animal cells in conditions of intensive agitation (1995)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 45 (1995), S. 463-472 
    Details
    ISSN: 0006-3592
    Keywords: apoptosis ; animal cell death ; hybridoma cells ; agitation ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The question is addressed as to whether cells which are subject to high-energy dissipation rates in agitated bioreactors show an apoptotic response. Murine hybridoma cells in batch culture were agitated in bench-scale (1-L) bioreactors without gas sparging. At an energy dissipation rate of 1.5 W m-3 there was no apparent damage. At 320 W m-3 cell viability declined, and increasing proportions of the dead cells displayed the morphological features of apoptosis, but necrosis also remained as a significant mechanism of death. When cells were subjected to the intensive energy dissipation rate of 1870 W m-3 in a bioreactor without gas headspace, the cell number dropped by 50% within 2 h and a subpopulation of smaller-sized cells emerged. This excluded trypan blue but showed some apoptotic characteristics such as reduced and condensed DNA content and low F-actin content. The incidence of apoptotic activity was further demonstrated by the appearance of numerous apoptotic bodies. Analysis of the cell cycles of both small and normal size populations indicated that greater proportions of S and G2 cells had become apoptotic and there was evidence of preferential survival of G1 cells. It is suggested that two mechanisms of cell death are apparent in hydrodynamically stressful situations, but their relative expression depends on the energy dissipation rate. © 1995 John Wiley & Sons, Inc.
    Additional Material: 9 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/bit.260450602
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  • 8
    Electronic Resource
    Electronic Resource
    Apoptosis and its control in cell culture systems (1997)
    Springer
    Cytotechnology 23 (1997), S. 87-93 
    Details
    ISSN: 1573-0778
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Apoptosis is a form of programmed cell death which exhibits highly distinctive morphology. Research activity in this area has increased substantially in recent years, primarily due to the realisation that disregulation of apoptosis is involved in the development of a number of pathological conditions, including cancer and AIDS. However, it is now clear that apoptosis also represents the dominant form of cell death during the culture of industrially important cell lines. This review focuses on the induction of apoptosis during industrial cell cultures as well as the effects of the apoptosis suppresser gene bcl-2 on cell survival in conditions relevant to bioreaction environments. We also present new data which demonstrates that bcl-2 can protect cells from apoptosis induced by oxygen deprivation, a finding which has important implications for large scale and intensive cultivation of cells. We also describe experiments which suggest that bcl-2 can reduce the specific nutrient consumption rate of cells.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007971703392
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  • 9
    Electronic Resource
    Electronic Resource
    Variable functions of bcl-2 in mediating bioreactor stress- induced apoptosis in hybridoma cells (1998)
    Springer
    Cytotechnology 28 (1998), S. 177-188 
    Details
    ISSN: 1573-0778
    Keywords: apoptosis ; bcl-2 ; cell death ; hybridoma ; osmolarity ; pH ; shear ; stress
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract It has been demonstrated that the cell lines used for production of biopharmaceuticals are highly susceptible to apoptosis, and that over-expression of the bcl-2 oncogene can protect cells from death. Stress associated with the deprivation of nutrients has been shown to be the main cause of apoptosis in culture. We have extended these studies by investigating the mechanism of cell death under conditions of sub-optimal pH, shear stress and hyperosmolarity, and the protective action of bcl-2 over-expression. At pH 6, there was no clear evidence of protection from cell death. However, at pH 8, the viability of the bcl-2 transfected cells was about 20% higher relative to the control cells. Cultivation of control cells in a flat bottomed bioreactor with a magnetic stirrer bar without a pivot ring resulted in exposure of the cells to a high attrition effect. As a result, cell growth was retarded and a high level of cell death by apoptosis was observed. Under the same conditions, the bcl-2 transfected cell line exhibited a nearly five fold increase in viable cell number. This finding indicates that under apoptosis-suppressed conditions, shear stress can stimulate cell growth. Batch cultivation of both control and bcl-2 transfected cells in 350 and 400 mOsm media resulted in suppression of cell growth, athough the effect was most marked in the control cell line. Adaptation of control cells to 400 mOsm proved to be impossible to achieve. However, the bcl-2 transfected cells exhibited resistance to the osmotic stress resulting in long term adaptation to a high salt environment. Specific productivity of bcl-2 transfected cells grown in high osmolarity medium was 100% higher than that produced by non- adapted bcl-2 transfected cells grown in normal osmolarity medium. These results demonstrate that bcl-2 has a beneficial effect on hybridoma cultivation under a wide range of culture stresses.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008002319400
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  • 10
    Electronic Resource
    Electronic Resource
    Influence of bcl-2 on antibody productivity in high cell density perfusion cultures of hybridoma (1999)
    Springer
    Cytotechnology 30 (1999), S. 95-106 
    Details
    ISSN: 1573-0778
    Keywords: apoptosis ; Bcl-2 ; fixed-bed ; hollow fibre ; hybridoma ; perfusion ; protein-free medium
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Apoptosis is an active, genetically determined death mechanism which can be induced by a wide range of physiological factors and by mild stress. It is the predominant form of cell death during the production of antibodies from murine hybridoma cell lines. A number of studies have now demonstrated that the suppression of this death pathway, by means of over-expression of survival genes such as bcl-2, results in improved cellular robustness and antibody productivity during batch culture. In the present study, the influence of bcl-2 expression on hybridoma productivity in two high density perfusion bioreactor systems was investigated. In the first system, a fixed-bed reactor, the DNA content in the spent medium was 25% higher in the control (TB/C3-pEF) culture than that found in the bcl-2 transfected (TB/C3-bcl2) cultures at all perfusion rates. This is indicative of a higher level of cell death in the control cell line. The average antibody concentration for the TB/C3-pEF cell line was 14.9 mg L-1 at perfusion rates of 2.6 and 5.2 d-1. However, for the TB/C3-bcl2 cell line it was 33 mg L-1 at dilution rates of 2 and 4 d-1. A substantial increase in antibody concentration was also found in the Integra Tecnomouse hollow fibre reactor. The antibody titre in the TB/C3-bcl2 cassette was nearly 100% higher than that in the TB/C3-pEF cassette during the cultivation period which lasted 6 weeks. Clearly, these results demonstrate the positive impact of bcl-2 over-expression on production of antibody in hybridoma perfusion cultures.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008055702079
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