Cell Growth and Protein Expression Kinetics

Kompala, Dhinakar S.

doi:10.1002/0471250570.spi039

Cited by 1 publication

(1 citation statement)

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“…We have previously demonstrated that the S phase SV40 promoter driving the expression of the intracellular reporter protein, β‐galactosidase, in host CHO‐DG44 cells demonstrates strong growth‐associated production (18). In addition, we have also shown that by replacing the SV40 promoter with a G1 phase specific AML promoter, the same host cells can be switched to inverse‐growth‐associated production kinetics (19, 20). While the cell cycle phase‐specific expression of the induced MMTV‐GR promoter system has not been characterized, the results presented here demonstrate inverse‐growth‐associated production of a secreted reporter glyco‐protein in recombinant CHO cells.…”

Section: Discussionmentioning

confidence: 93%

Production of a Secreted Glycoprotein from an Inducible Promoter System in a Perfusion Bioreactor

2004

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The primary advantage of an inducible promoter expression system is that production of the recombinant protein can be biochemically controlled, allowing for the separation of unique growth and production phases of the culture. During the growth phase, the culture is rapidly grown to high cell density prior to induction without the extra metabolic burden of exogenous protein production, thus minimizing the nonproductive period of the culture. Induction of the culture at high cell density ensures that the volumetric production will be maximized. In this work, we have demonstrated the feasibility of overexpressing a reporter glycoprotein from the inducible MMTV promoter in recombinant Chinese hamster ovary (CHO) cells cultured in a high cell density perfusion bioreactor system. Retention of suspension-adapted CHO cells was achieved by inclined sedimentation. To maximize volumetric production of the culture, we have demonstrated that high cell density must be achieved prior to induction. This operating scheme resulted in a 10-fold increase in volumetric titer over the low density induction culture, corresponding directly to a 10-fold increase in viable cell density during the highly productive period of the culture. The amount of glycoprotein produced in this high cell density induction culture during 26 days was 84-fold greater than that produced in a week long batch bioreactor. Long-term perfusion cultures of the recombinant cell line showed a production instability, a phenomenon that is currently being investigated.

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Section: Discussionmentioning

confidence: 93%