Rational scale-up of a baculovirus-insect cell batch process based on medium nutritional depth

Yang, Jeng‐Dar; Gecik, Peter; Collins, Alaric; Czarnecki, Stephen; Hsu, Hong‐Hue; Lasdun, Abraham; Sundaram, Ramakrishnan; Muthukumar, Ganapathy; Silberklang, Melvin

doi:10.1002/(sici)1097-0290(19961220)52:6<696::aid-bit7>3.0.co;2-j

Cited by 30 publications

(32 citation statements)

References 30 publications

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“…The negative relation was not only for the late stage of recombinant protein production but also for the earlier stages of vDNA replication, extracellular vDNA production and mRNA expression. This finding is the main contribution of this work compared with previous studies that only reported the negative relation between the PCD and recombinant protein yields for Hi5/ AcMNPV infections (Chico and Jager 2000;Ikonomou et al 2004;Taticek and Shuler 1997;Yang et al 1996). In addition, the finding from the current study together with the previous study with the Sf9/rAcMNPV system (Huynh et al 2013) confirms that different insect cell lines infected with a recombinant baculovirus generate similar outcomes in their response to the increase in the PCD.…”

Section: Discussionsupporting

confidence: 81%

“…Thus, the lack of a stable region for specific yields above the PCD of 0.6×10 6 cells/mL for infected Hi5 cells may be explained by this cell size difference. A reduction in the specific recombinant protein yield for infected T. ni cells above a PCD of 0.6×10 6 cells/mL was also reported in ExCell 401 and TNM-FH media (Chico and Jager 2000;Taticek and Shuler 1997), while others reported a stable region for infections over the range of 0.5-2×10 6 cells/mL in Ex-Cell 405 medium (Yang et al 1996). Fig.…”

Section: Discussionmentioning

confidence: 61%

“…Significance testing was performed separately for each experimental set. Data points with different letters are significantly different according to Tukey's HSD (α=0.05) ammonia to toxic levels (Rhiel et al 1997;Sugiura and Amann 1996;Yang et al 1996), whereas, Sf9 cell lines are not associated with extensive lactate or ammonium accumulation (Bedard et al 1993;Benslimane et al 2005;Drews et al 2000;Rhiel et al 1997;Sugiura and Amann 1996). Any possible toxin-induced cell death effects were less severe in the case of fresh medium infections (Fig.…”

Section: Discussionmentioning

confidence: 95%

“…This study is useful in order to clarify whether the above phenomenon is cell line specific or likely to be a general behaviour of baculovirus-infected insect cell systems. Although a cell-specific yield reduction with increasing ICDs has been reported previously for the T. ni cell line (Chico and Jager 2000;Ikonomou et al 2004;Taticek and Shuler 1997;Yang et al 1996), these early studies were not designed for investigating yields over a wide range of ICDs and did not involve the up-stream analysis of virus replication and transcription to understand whether the cell density effect was a late infection phenomenon or was the result of early limitations in virus replication and mRNA expression.…”

Section: Discussionmentioning

confidence: 99%

“…The so-called cell density effect has been reported thoroughly for Spodoptera frugiperda (Sf9/S921) cell lines (Carinhas et al 2009;Caron et al 1990;Doverskog et al 2000;Huynh et al 2013;Jesionowski and Ataai 1997;Radford et al 1997;Reuveny et al 1993;Taticek and Shuler 1997;Wong et al 1996;Yamaji et al 1999) and to a lesser extent for the Trichoplusia ni (Hi5) cell line (Chico and Jager 2000;Ikonomou et al 2004;Taticek and Shuler 1997;Yang et al 1996). This issue is a key hurdle for improving the yield of the baculovirus-infected insect cell systems in an attempt to reduce production costs.…”

Section: Introductionmentioning

confidence: 99%

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Effect of the peak cell density of recombinant AcMNPV-infected Hi5 cells on baculovirus yields

Huynh

Tran

Chan

et al. 2014

Appl Microbiol Biotechnol

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The phenomenon of the cell density effect is not readily explained by an obvious nutrient limitation, and a recent study has suggested that for recombinant Autographa californica multiple nucleopolyhedrovirus (rAcMNPV)-infected Sf9 cells, a drop in messenger RNA (mRNA) levels may be sufficient to explain the cell density effect for this system. The current study aims to investigate the response in cell-specific yields (viral DNA (vDNA), LacZ mRNA and β-galactosidase (β-Gal) protein) with increasing infection cell density (ICD) for rAcMNPV-infected Hi5 cells, where the rAcMNPV expresses the β-Gal gene under control of the polyhedral promoter. Hi5 cells in suspension culture of Express Five® medium were synchronously infected with a rAcMNPV at multiple ICDs between 0.5 and 6 × 10(6) cells/mL and a multiplicity of infection of 10 plaque-forming units (PFU)/cell either in the original or fresh medium conditions. There were negative correlations between the three key virus infection indicators (vDNA, mRNA and β-Gal) and the peak cell density (PCD). However, unlike infected Sf9 cells, the yield decline started at the lowest PCD investigated (0.6 × 10(6) cells/mL). Generally, the yield decline with increasing PCD was most pronounced for β-Gal followed by mRNA and was more moderate for vDNA. The decline was significantly reduced but not totally arrested when fresh medium replacement was used. The results suggest that the reduction in recombinant protein-specific yields at high PCDs is associated with limitations during the up-stream processes of replication and transcription rather than entirely caused by limitations during translation. In addition, low production rates at late infection stages of moderate to high ICDs are a probable cause of the cell density effect.

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Section: Discussionsupporting

confidence: 81%

Section: Discussionmentioning

confidence: 61%

Section: Discussionmentioning

confidence: 95%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Effect of the peak cell density of recombinant AcMNPV-infected Hi5 cells on baculovirus yields

Huynh

Tran

Chan

et al. 2014

Appl Microbiol Biotechnol

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Glycosylation of a recombinant protein in the Tn5B1-4 insect cell line: Influence of ammonia, time of harvest, temperature, and dissolved oxygen

Donaldson

Wood

Kulakosky

et al. 1999

Biotechnol. Bioeng.

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Glycosylation is both cell line and protein dependent. Culture conditions can also influence the profile of glycoforms produced. To examine this possibility in the insect cell/baculovirus system, structures of N‐linked oligosaccharides attached to SEAP (human secreted alkaline phosphatase), expressed under various culture conditions in BTI Tn5B1‐4 cells, were characterized using FACE (fluorescence‐assisted carbohydrate electrophoresis). Parameters varied were time of harvest, ammonia added during infection, dissolved oxygen, and temperature. It was found that glycosylation in the insect cell/baculovirus expression system is a robust, stable system that is less perturbed by variations in culture conditions than the level of protein expression. Addition of ammonia and low oxygen conditions affected SEAP expression, but not the oligosaccharide profile of SEAP. Time of SEAP harvest increased the amount of α‐mannosidase resistant structures from 4.1% at 34 hours postinfection (h pi), to 5.0% at 100 h pi, and to 7.5% at 120 h pi. These structures were primarily sensitive to N‐acetylhexosaminidase digest, although a small amount was insensitive to both mannosidase and N‐acetyl‐hexosaminidase digests. Lowering the temperature from 28°C to 24°C or even 20°C, resulted in a twofold increase in oligosaccharides containing terminal α(1,3)‐mannose residues. This condition did not affect the amount of mannosidase‐resistant structures. However, this could result in more complete glycosylation of recombinant proteins in the BTI Tn5B1‐4 cell line, because more structures with the potential for further processing would be produced. © 1999 John Wiley & Sons, Inc. Biotechnol Bioeng 63: 255–262, 1999.

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Optimized production of recombinant bluetongue core-like particles produced by the baculovirus expression system

Zheng

Greenfield

Reid

1999

Biotechnol. Bioeng.

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The baculovirus‐expression vector system (BEVS) has been widely used for the experimental production of many human and animal single‐ and multi‐unit vaccines, heterologous proteins, and viral insecticides. In this work, the production of recombinant bluetongue virus core‐like particles (CLPs), using Sf9 cells in shaker‐suspension culture with the SF900 II medium (GIBCO, NY), has been studied. This system involved the simultaneous production of two proteins, VP7 and VP3, and was shown to achieve high volumetric productivities. The key parameters of the time of infection (TOI), and the multiplicity of infection (MOI) were studied. The results show that the peak‐volumetric yields and cell‐specific yields achieved using low MOIs at low‐cell densities were the same as those obtained following infections with a high MOI at high‐cell densities. This work establishes the feasibility of using low MOIs in the baculovirus system to produce complex multiprotein particles. © 1999 John Wiley & Sons, Inc. Biotechnol Bioeng 65: 600–604, 1999.

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Rational scale-up of a baculovirus-insect cell batch process based on medium nutritional depth

Cited by 30 publications

References 30 publications

Effect of the peak cell density of recombinant AcMNPV-infected Hi5 cells on baculovirus yields

Effect of the peak cell density of recombinant AcMNPV-infected Hi5 cells on baculovirus yields

Glycosylation of a recombinant protein in the Tn5B1-4 insect cell line: Influence of ammonia, time of harvest, temperature, and dissolved oxygen

Optimized production of recombinant bluetongue core-like particles produced by the baculovirus expression system

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