• Title/Summary/Keyword: specific monoclonal antibody productivity

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Monoclonal Antibody Refolding and Assembly: Protein Disulfide Isomerase Reaction Kinetics

  • Park, Sun-Ho;Ryu, Dewey D.Y.
    • Biotechnology and Bioprocess Engineering:BBE
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    • v.8 no.2
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    • pp.59-63
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    • 2003
  • The protein disulfide isomerase (PDI) reaction kinetics has been studied to evaluate its effect on the monoclonal antibody (Mab) refolding and assembly which accompanies disulfide bend formation. The MAb in vitro assembly experiments showed that the assembly rate of heavy and light chains can be greatly enhanced in the presence of PDI as compared to the rate of assembly obtained by the air-oxidation. The reassembly patterns of MAb in-termediates were identical for both with and without PDI, suggesting that the PDI does not determine the MAb assembly pathway, but rather facilitates the rate of MAb assembly by promoting PDI catalyzed disulfide bond formation. The effect of growth rate on PDI activities for MAb production has also been examined by using continuous culture system. The specific MAb productivity of hybridoma cells decreased as the growth rate increased. However, PDI activities were nearly constant fur a wide range of growth rates except very high growth rate, indicating that no direct correlation between PDI activity and specific MAb productivity exists.

Monoclonal Antibody Refolding and Assembly: Protein Disulfide Isomerase Reaction Kinetics

  • Park, Sun-Ho;Ryu, Dewey D.Y.
    • Biotechnology and Bioprocess Engineering:BBE
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    • v.1 no.1
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    • pp.13-17
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    • 1996
  • The protein disulfide isomerase(PDI) reaction kinetics has been studied to evaluate its effect on the monoclonal antibody(MAb) refolding and assembly which accompanies disulfide bond formation The MAb in vitro assembly experiments showed that the assembly rate of heavy and light chains can be greatly enhanced in the presence of PDI as compared to the rate of assembly obtained by the air-oxidation. The reassembly patterns of MAb intermediates were identical for both with and without PDI, suggesting that the PDI does not determine the MAb assembly pathway, but rather facilitates the rate of MAb assembly by promoting PDI catalyzed disulfide bond formation. The effect of growth rate on PDI activities for MAb production has also been examined by using continuous culture system. The specific MAb productivity of hybridoma cells decreased as the growth rate increased. However, PDI activities were nearly constant for a wide range of growth rates except very high growth rate, indicating that no direct correlation between PDI activity and specific MAb productivity exists.

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Effect of Intracellular Calcium Level on the Hybridoma Cell Growth and Monoclonal Antibody Production (세포내 calcuim 농도가 하이브리도마 세포 성장 및 단일클론항체 생산에 미치는 영향)

  • 박재성;남민희;박선호
    • KSBB Journal
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    • v.13 no.5
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    • pp.585-592
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    • 1998
  • The effect of intracellular Ca2+ level on the hybridoma cell growth and monoclonal antibody(MAb) production was examined. For the manipulation of intracellular Ca2+ concentration, the cells were treated with A23187, ryanodine, and thapsigargin at about 1x106 cells/mL. The treated cells were recultivated by using the Iscove's Modified Dulbecco's Medium(MDM) containing 1.49mM CaCl2. The ryanodine-treated cells showed better cell growth, MAb concentration, and specific MAb productivity than others. In comparison with control, the maximum cell concentration, MAb concentration, and specific MAb productivity were increased by 40.6%, 48.1% and 83.3%, respectively. Confocal microscopic images of Fura-2/AM loaded cells indicate that the increase in intracellular Ca2+ level can enhance the MAb productivity by allowing the calcium influx into the endoplasmic reticulumn.

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The Effect of Media Feeding Rate on the Production of Monoclonal Antibody Production in the Fed-batch Culture of Hybridoma (하이브리도마 세포의 유가식 배양에서 배지첨가속도가 단일클론 항체 생산에 미치는 영향)

  • 곽원재;최태부;박정극
    • Microbiology and Biotechnology Letters
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    • v.19 no.3
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    • pp.272-280
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    • 1991
  • The effect of media feeding rate on cell growth and monoclonal antibody production in the fed-batch culture ot hybridoma A4W was studied. In the batch culture, the highest specific antibody production rate was observed at the begining of the culture period but its value tended to decrease rapidly with the culture time. The final antibody concentration and volumetric productivity was 65 $\mu g$/ml and 13 mg Mab/l/day, respectively. In the fed-batch culture, the specific antibody production rate, $q_p$ rebounded sharply within a few hours after the media feeding was started and it remained high until the end of culture if the media feeding was continued. The final antibody concentration was 220 $\mu g$/ml and the volumetric productivity was 45.1 mg/l/day. Further increase in final antibody concentration was achieved by applying a modified media of which component was fortified with glucose and glutamine, hence the final antibody concentration in this case was 270 $\mu g$/ml and the volumetric productivity was 51.8 mg/lday, which is as four tinlcs as high cuixparinf! to that of batch culture.

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Effects of high Cell Density on growth-Associated Monoclonal Antibody Production by Hybridoma T0405 Cells Immobilized in Macroporous Cellulose carriers

  • Hideki Mochoda;Wang, Pi-Chao;Fr Jr. Nayve;Ryuji Sato;Minoru Harige;Nakao Nomura;Masatoshi Matsumura
    • Biotechnology and Bioprocess Engineering:BBE
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    • v.5 no.2
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    • pp.110-117
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    • 2000
  • Relationship between monoclonal antibody (MAb) productivity and growth rate, and effects of high cell density on MAb production rate increased with increasing specifis growth rate in both suspended and immobilized continuous cultures indicate a positively growth-associated relationship between MAb productivity and growth rate. moreover, the specific production rate was higher in the immobilized cell culture than that in suspended one at all dilution rates. In order to clarify these phenomana, MAb mPNA experession and cell cycle distribution were investigated in bacth cultures with immobilized cells and suspended cells. RT-PCR was used for observation of MAb mRNA expression and a two-color bromodeoxyuridine (BrdU)/propidium iodide (PI) flow cytometry method for determination of cell cycle distribution. The results revealed that MAb nRNA expression until dead phase, which was longer than in suspended cell. The cell cycle distribution patterns were observed almost the same for both immobilized and suspended cells. Such results may imply that a high cell density state has positive influence on the mRNA expression and on growth-associated Mab productivity of T0405 cells.

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Glucose Effects on Cell Growth, Antibody Production, and Cell Metabolism of Hybridoma Cells (Hybridoma 세포의 세포성장, 항체생산 및 세포대사에 미치는 Glucose의 영향)

  • ;Shaw S.Wang
    • KSBB Journal
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    • v.10 no.3
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    • pp.323-334
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    • 1995
  • The effects of glucose on cell growth kinetics, monoclonal antibody productivity, and cell metabolism or hybridoma cells were investigated. The mouse-mouse hybridoma cell line VIII H-8 producing mouse IgG2a was used as a modal system. Glucose showed substrate inhibition type dependence on specific growth raie. The maximum cell density increased as initial glucose concentration increased up to 4 g/$\ell$. Glucose showed a strong influence on cell death kinetics, and an inverse relationship between specific death rate and glucose concentration was found. Cell viability and monoclonal antibody production increased as initial glucose concentration increased. The specific glucose consumption rate increased with glucose concentration, and cumulative specific lactate production rate increased with increasing initial glucose concentration. The overall kinetics of ammonium ion production was almost invariant with respect to initial glucose concentration, while the cumulative specific ammonium ion production rate was dependent on initial glucose concentration.

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Modelling of a Biomolecular Processing for the Production and Secretion of Monoclounal Antibody (단일콜론항체 생산 및 분비에 대한 생물분자공정의 모델링)

  • 박재성;박선호
    • KSBB Journal
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    • v.13 no.4
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    • pp.369-377
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    • 1998
  • To analyze the unique aspects of biomolecular processing for monoclonal antibody (MAb) production and secretion, the simple working model based on 3-compartment (endoplasmic reticulum, Golgi apparatus, and extracellular medium) was developed. Based on in vitro MAb assembly experimental results, the kinetic model for MAb assembly in the endoplasmic reticulumn was proposed. The dynamics of MAb assembly and secretion was simulated using methematica program. According to the simulation results, the proposed 3-compartment model provides an efficient means to predict the specific MAb productivity as well as intracompartmental concentrations of MAb in endoplasmic reticulum, Golgi apparatus, and extracellular compartment model. In vivo profiles of MAb intermediates gave good agreements with the simulation profiles predicted by the intracellular compartment model. Furthermore, results of such analysis can help in directing the control strategy for optimum biomolecular processing in a mammalian cell culture system.

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Characterization of Physiological Changes in $S3H5/\gamma{2bA2}$ Hybridoma Cells During Adaptation to Low Serum Media

  • Lee, Gyun-Min;Joanne, Savinell
    • Journal of Microbiology and Biotechnology
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    • v.2 no.2
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    • pp.141-151
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    • 1992
  • Physiological changes of the murine hybridoma cell line $S3H5/\gamma{2bA2}$ during adaptation to RPMI 1640 medium with 1%(v/v) fetal bovine serum were characterized in terms of cell growth, antibody production, morphology, and metabolic quotients. Cells adapted to 1% serum medium in T-flasks became sensitive to shear induced by mechanical agitation and required at least 5% serum in the medium or spent medium for cell growth in spinner flasks, while cells adapted to 10% serum medium in T-flasks could grow in 1% serum medium in spinner flasks. Consequently, long-term adaptation to low serum media may not give the expected growth enhancement. After adaptation to 1% serum medium, changes in cell morphology were observed. The cells in 10% serum medium were uniform and circular, while cells in 1% medium were irregularly shaped. The DNA contents, which were measured by flow cytometry, were almost constant among the cells in the range of 1% to 10%. Further, no significant changes in energy metabolism and specific monoclonal antibody production rate were observed among these cells.

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Effects of Glucose and Acetic Acid on the Growth of Recombinant E.coli and the Production of Pyruvate Dehydrogenase Complex-E2 Specific Human Monoclonal Antibody (유전자 재조합 대장균의 세포성장과 Pyruvate Dehydrogenase Complex-E2 특이성 인간 모노클론 항체 생산에 대한 포도당과 초산의 영향)

  • 이미숙;전주미;차상훈;정연호
    • KSBB Journal
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    • v.15 no.5
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    • pp.482-488
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    • 2000
  • The Fab fraction of PDC-E2 specific human monoclonal antibody was produced using recombinant E. coli, and the effects of glucose and acetate were investigated to develop an optimal strategy for recombinant human antibody production. Higher glucose concentration in the culture media resulted inn higher cell growth and glucose consumption rate, which in turn resulted in an increased acetate production rate. When glucose was depleted, cells began to consume acetate as an energy source, and this consumption rate depended on the glucose concentration. When the residual glucose concentration was high, the accumulation of acetate was accelerated due to an increase in the acetate production rate and a decrease in the acetate consumption rate. Futhermore, it was found that a high accumulation of acetate, accompanied by a high glucose concentration, inhibited human antibody formation; the critical acetate concentration was $0.6g/\ell$. During production, a high glucose concentration enhanced cell growth, but inhibited antibody formation due to catabolic repression. Therefore, it is important to keep the concentration of both glucose and acetate as low as possible to increase antibody production after induction. Accordingly, it is important to accurately control the concentration of glucose and acetate in the culture media to obtain high cell densities and high productivity levels of recombinant human antibody.

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High Density Culture of KA112 Hybridoma and Effect of Glucose Concentration on MAb Productivity (하이브리도마의 고농도 배양과 포도당 농도가 MAb 생산성에 미치는 영향)

  • 박상재;최차용
    • KSBB Journal
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    • v.8 no.5
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    • pp.478-482
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    • 1993
  • Perfusion culture was conducted in Celligen perfusion culture system using a self-constructed hybridoma cell and low serum medium. The culture system employed hollow fiber to separate cells from the culture broth. Maximum cell density of $2.1\times10^7$ ce11s/m1, 10 times higher than in batch culture, could be achieved. Concentration of monoclonal antibody (MAb) was 4 times higher and production rate at maximum feed rate was 9 times higher than in batch culture. Glucose concentration was very important for the cell growth and MAb production. When glucose concentration was below 1g/l, i. e. 0.5~0.9g/l, specific MAb production rate decreased but cell concentration still increased. As the glucose concentration goes above 1g/l, specific MAb production rate increased and remained at maximum value at more than 1.5g glucose/l. The maximum value of the specific Mab production rate was similar to that of batch culture.

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