Perfusion culture of hybridoma cells using a centrifuge to separate cells from culture mixture

Hamamoto, Kimihiko; Ishimaru, Kenji; Tokashiki, Michiyuki

doi:10.1016/0922-338x(89)90121-9

Cited by 38 publications

(8 citation statements)

References 8 publications

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“…Intermittent-centrifugation perfusion was evaluated to increase the fraction of supernatant extracted from the feed and reduce the frequency of cell passage to the centrifuge, this under the hypothesis that cell viability or productivity may be less affected by longer but occasional oxygen limitations than by the frequent but shorter ones of continuous procedure. Other authors (Hamamoto et al, 1989;Tokashiki et al, 1990) have shown that X87 and H1 hybridomas in batch culture submitted to 10-30-min centrifugations at 100g twice a day exhibited growth and MAb production similar to those of control cultures. In the experiments reported here, while fresh medium was fed continuously to the bioreactor, the cell suspension was circulated through the centrifuge only twice a day, for 30 min each time.…”

Section: Resultsmentioning

confidence: 80%

“…Also, with the higher culture residence time inside the bioreactor, the cells experienced less shear stress and an overall shorter time under nutrient limitation in the insert bottom. Other authors (Hamamoto et al, 1989; Tokashiki et al, 1990), who described the growth of a mouse‐human X87 hybridoma producing an IgG1, found that production was equivalent using either continuous centrifugation, intermittent centrifugation, or gravitational settling. However, 4‐ and 10‐L bioreactors were used in those studies, which may not be comparable to 1.2−1.3‐L vessels, since the cell‐passage frequency is likely to be lower with larger reactors.…”

Section: Resultsmentioning

confidence: 99%

“…Other continuous centrifuge models have been used at up to 63g without affecting cell growth or MAb productivity (Tokashiki et al, 1990). It has also been reported that cells can be subjected to up to 200g without a loss of viability and productivity (Hamamoto et al, 1989;Shimazaki et al, 1986).…”

Section: Resultsmentioning

confidence: 99%

“…We have assessed the utility of alternative continuous cell‐retention devices for the long‐term culture of hybridoma cells. The acoustic aggregation sedimenter (Gaida et al, 1996; Kilburn et al, 1989; Trampler et al, 1994) and centrifugal separator (Apelman, 1991; Apelman and Björling, 1991; Hamamoto et al, 1989; Hodgson, 1991; Tokashiki et al, 1990; Tokashiki and Takamatsu, 1993) were identified as viable techniques, exploiting the inertial properties of the cells instead of using physical barriers, to perform the separation. This paper focuses on the evaluation of a bench‐top centrifugal separator, the first prototype of the Centritech Lab (Centritech AB, Norsborg, Sweden).…”

Section: Introductionmentioning

confidence: 99%

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Use of the Centritech Lab Centrifuge for Perfusion Culture of Hybridoma Cells in Protein‐Free Medium

Johnson¹,

Lanthier²,

Massie³

et al. 1996

Biotechnology Progress

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As part of an effort to develop a suspension-culture perfusion-based process with high flow rate without the fouling and antibody retention inherent to filter-based cell-separation devices, we have evaluated and contributed to the development of the Centritech Lab centrifuge for the perfusion culture of hybridoma cells in protein-free medium. Culture start-ups showed that cell growth and monoclonal-antibody (MAb) production rates were similar in both a spinner flask and continuous centrifugation coupled to a bioreactor. The centrifuge efficiently separated viable cells from dead ones. Viable-cell recoveries were never below 98%, whereas dead-cell recoveries were usually around 80%. The cell content of the centrifuge supernatant and concentrate was strongly determined by the total amount of cells, viable and dead, in the culture broth, but an influence of the centrifugation parameters (feed rate, times of separation and discharge, and rotor speed) was observed. This understanding of the separation process inside the centrifuge is important and may apply to other similar devices. Monoclonal antibodies were not retained in the bioreactor during centrifugation perfusion. However, whereas similar growth rates were obtained in perfusion cultures using either continuous centrifugation or filtration, MAb concentrations were 35% lower in the former case. Utilization of the centrifuge in an intermittent fashion decreased the daily cell residence time outside the bioreactor, the daily pelleted-cell residence time in the centrifuge, and the frequency of cell passage to the centrifuge. This led to higher viable-cell numbers in the bioreactor and an accompanying increase in MAb concentrations, 225-250 mg of IgM L-1, equal to the performance of filter-based perfusion systems with the same cell line. It was hypothesized that having cells periodically packed at the bottom of the centrifuge insert (up to 800 x 10(6) cells mL-1) is deleterious to the culture by exposing the pelleted cells to prolonged nutrient limitations.

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Section: Resultsmentioning

confidence: 80%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Use of the Centritech Lab Centrifuge for Perfusion Culture of Hybridoma Cells in Protein‐Free Medium

Johnson¹,

Lanthier²,

Massie³

et al. 1996

Biotechnology Progress

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show abstract

“…Several investigators designed custom centrifuges for perfusion culture, working either continuously (Tokashiki et al, 1990) or semicontinuously (Hamamoto et al, 1989;Takamatsu et al, 1996). During the perfusion cultures using a centrifuge for cell retention, no report was made concerning a detrimental effect of centrifugation on animal cells.…”

Section: Voisard Et Al: Large-scale Retention Of Mammalian Cellsmentioning

confidence: 98%

Potential of cell retention techniques for large‐scale high‐density perfusion culture of suspended mammalian cells

Voisard

Meuwly

Ruffieux

et al. 2003

Biotech & Bioengineering

244

150

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This review focuses on cultivation of mammalian cells in a suspended perfusion mode. The major technological limitation in the scaling-up of these systems is the need for robust retention devices to enable perfusion of medium as needed. For this, cell retention techniques available to date are presented, namely, cross-flow filters, hollow fibers, controlled-shear filters, vortex-flow filters, spin-filters, gravity settlers, centrifuges, acoustic settlers, and hydrocyclones. These retention techniques are compared and evaluated for their respective advantages and potential for large-scale utilization in the context of industrial manufacturing processes. This analysis shows certain techniques have a limited range of perfusion rate where they can be implemented (most microfiltration techniques). On the other hand, techniques were identified that have shown high perfusion capacity (centrifuges and spin-filters), or have a good potential for scale-up (acoustic settlers and inclined settlers). The literature clearly shows that reasonable solutions exist to develop large-scale perfusion processes.

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Stirred Tank Reactors for Cell Culture Technology

Pangarkar

2014

Design of Multiphase Reactors

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Perfusion culture of hybridoma cells using a centrifuge to separate cells from culture mixture

Cited by 38 publications

References 8 publications

Use of the Centritech Lab Centrifuge for Perfusion Culture of Hybridoma Cells in Protein‐Free Medium

Use of the Centritech Lab Centrifuge for Perfusion Culture of Hybridoma Cells in Protein‐Free Medium

Potential of cell retention techniques for large‐scale high‐density perfusion culture of suspended mammalian cells

Stirred Tank Reactors for Cell Culture Technology

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