Methods for increasing monoclonal antibody production in suspension and entrapped cell cultures: biochemical and flow cytometric analysis as a function of medium serum content

Heath, Carole A.; Dilwith, Robert L.; Belfort, Georges

doi:10.1016/0168-1656(90)90052-d

Cited by 58 publications

(38 citation statements)

References 18 publications

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“…However, they also prove that the presence of serum in the culture medium, even when added to the SFM, has pronounced inhibitory effect on cell specific MAB productivity. Similar results, that the omission or reduction of serum in the culture medium leads to enhanced MAB production per cell, have been reported by Chang et al (1980), Chua et al (1994), Heath et al (1990), Mariani et al (1991) and .…”

Section: Medium Effectssupporting

confidence: 87%

Effect of serum-free and serum-containing medium on cellular levels of ER-based proteins in various mouse hybridoma cell lines

Lambert

Merten²

1997

Biotechnol. Bioeng.

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Section: Medium Effectssupporting

confidence: 87%

Effect of serum-free and serum-containing medium on cellular levels of ER-based proteins in various mouse hybridoma cell lines

Lambert

Merten²

1997

Biotechnol. Bioeng.

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“…The monoclonal antibody produced was cell growth associated, a kinetic type often observed [19]. The rate Y LAC/GLU (0.69 g lactate/ g glucose) also ®ts the literature data for hybridomas [12].…”

Section: Discussionmentioning

confidence: 61%

Glucose uptake rate as a tool to estimate hybridoma growth in a packed bed bioreactor

Rodrigues

Vilaça

Garbuio

et al. 1999

Bioprocess Engineering

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The immobilisation of cells in a perfusion culture allows to obtain a high cell concentration and an ef®cient removal of the catabolites without cell loss. A disadvantage of this system is that the cell density cannot be directly monitored. The cellular metabolism is just followed by online measurements of pH and dissolved oxygen (DO) and off-line determinations of residual metabolites. In this article, we report a high cell density achieved by the cultivation of a hybridoma in a bubblecolumn bioreactor ®lled with hollow glass cylinders. The parameters monitored during the cultivation were pH, temperature, DO, glucose, lactate and monoclonal antibody. The glucose uptake rate was used to estimate the cell concentration along the time. The maximum cell concentration calculated for the considered cultivation time was 2.7´10 7 cell á ml A1 . The glucose concentration in the media decreased stepwise twice, causing a decrease on the speci®c growth rate, while maintaining high antibody productivity levels. Maximum monoclonal antibody productivity was 503 lg á l A1 á day A1 and speci®c productivity, considering calculated cell density, was 0.019 ng á cell A1 á day A1 . IntroductionAlthough target of a great number of studies, mammalian cell culture kinetics is still not completely understood. It depends on so many factors, including the cell line physiology, media formulation, type of bioreactor, gas supply and operating conditions. Often, the particular characteristics of a given cell line are functions of the culture conditions, including the composition of the medium, the bioreactor type, and the way it is operated [1].The most commonly used basal media for the growth of hybridomas are DME and RPMI 1640, formulated with different concentrations of glucose and glutamine, the major nutrients related to growth in conventional cell culture media, representing cell mass precursors and energy source. The other nutrients do not count for growth limitation, as they are usually supplied in excess. In a comparison between DME and RPMI media, it was concluded that DME seems to be more ef®cient for hybridoma growth, supporting greater number of cells for longer periods than RPMI, while the antibody synthesis depends only on the cell number, regardless of the medium used [2,3].Concerning the culture mode, perfusion culture of mammalian cells allows the maintenance of long-term, high-density and high-productivity cultures. Suspension cultures of hybridoma in a perfusion system makes it dif®cult to control the problems caused by the shear forces and to increase the dilution rate above the maximum speci®c growth rate, permitting to achieve high cellular viability. Several devices have been attempted to retain the cells, with more or less ef®cacy [4±6]. The immobilisation of hybridoma cells in any kind of support allows more ef®cient perfusion and confers more stability to the culture system, therefore, increasing the potential for longer cultures, higher cell density and antibody productivity [7]. The immobilisation condition protect...

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“…Although apoptosis plays a role, necrosis may also be caused by extreme conditions. But often in bioreactors an environmental factor contributing to cell death steadily builds up, thus, termination of bioreactor cell cultures is often due to apoptosis rather than necrosis as a result of external factors such as excessive shear forces (Chisti 1993;Emery et al 1995;Kuystermans and Al-Rubeai 2011;Petersen et al 1988), metabolic nutrient depletion (Bibila et al 1994;Heath et al 1990;Leelavatcharamas et al 1994;Singh and AlRubeai 1998), metabolic by-product accumulation (Martinelle et al 1996;Xie and Wang 1994), hypoxia (Jan et al 1997;Ogawa et al 1992), sub-optimal pH (Naciri et al 2008;Singh 1994) and hyper-osmolality (deZengotita et al 2002). Since apoptosis is regulated through a cascade of molecular events in response to external factors (Arden and Betenbaugh 2004) it is possible to monitor the progress of apoptosis within a cell population.…”

Section: Introductionmentioning

confidence: 99%

Online flow cytometry for monitoring apoptosis in mammalian cell cultures as an application for process analytical technology

2014

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Apoptosis is the main driver of cell death in bioreactor suspension cell cultures during the production of biopharmaceuticals from animal cell lines. It is known that apoptosis also has an effect on the quality and quantity of the expressed recombinant protein. This has raised the importance of studying apoptosis for implementing culture optimization strategies. The work here describes a novel approach to obtain near real time data on proportion of viable, early apoptotic, late apoptotic and necrotic cell populations in a suspension CHO culture using automated sample preparation in conjunction with flow cytometry. The resultant online flow cytometry data can track the progression of apoptotic events in culture, aligning with analogous manual methodologies and giving similar results. The obtained near-real time apoptosis data are a significant improvement in monitoring capabilities and can lead to improved control strategies and research data on complex biological systems in bioreactor cultures in both academic and industrial settings focused on process analytical technology applications.

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Methods for increasing monoclonal antibody production in suspension and entrapped cell cultures: biochemical and flow cytometric analysis as a function of medium serum content

Cited by 58 publications

References 18 publications

Effect of serum-free and serum-containing medium on cellular levels of ER-based proteins in various mouse hybridoma cell lines

Effect of serum-free and serum-containing medium on cellular levels of ER-based proteins in various mouse hybridoma cell lines

Glucose uptake rate as a tool to estimate hybridoma growth in a packed bed bioreactor

Online flow cytometry for monitoring apoptosis in mammalian cell cultures as an application for process analytical technology

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