Chemically defined medium for the production of biologically active substances of CHO cells

Hata, Jun-ichiro; Tamura, Takeyoshi; Yokoshima, Soji; Yamashita, Shinya; Kabeno, Shoko; Matsumoto, Ken; Onodera, Kônoshin

doi:10.1007/bf00376095

Cited by 11 publications

(5 citation statements)

References 14 publications

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“…RBCF-1 cells, derived from goldfish fin [22], were propagated as a monolayer culture at 37°C in Dulbecco's modified Eagle's (Gibco):-Ham F12 (1:1) medium supplemented with 10% FCS [23]. A mixture of 0.01 Ixg of receptor plasmid (pCMV-rtGR-I or II) and 0.2 ~tg of reporter plasmid (pMSG-CAT) were cotransfected into subconfluent cultures of RBCF-1 cells in each well of 6-well dishes (Falcon) using calcium phosphate coprecipitation [24,25].…”

Section: Transient Gr Expression In Cultured Cellsmentioning

confidence: 99%

Fish glucocorticoid receptor with splicing variants in the DNA binding domain

et al. 1996

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Here we describe the isolation of a rainbow trout cDNA containing an entire GR coding region. Although the encoded protein is highly homologous to other GRs, especially in its DNA binding domain, it contains a nine amino acid insertion between the two zinc fingers. This novel form is found in all rainbow trout tissues examined; however, the testis also contains a splice variant lacking this insert, making it completely continuous to other GRs. In transient transfection assays of cultured cells, the two rainbow trout GR variants activated transcription from the glucocorticoid-responsive mouse mammary tumor virus promoter to comparable levels.

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Section: Transient Gr Expression In Cultured Cellsmentioning

confidence: 99%

Fish glucocorticoid receptor with splicing variants in the DNA binding domain

et al. 1996

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“…Development of various chemically defined media for NS0 cells and CHO cells has been reported (Chen et al, 2000;Hata et al, 1992;Huang et al, 2007;Zhang and Robinson, 2005), and commercial chemically defined media are readily available from all major mammalian cell culture medium manufacturers. However, the development of chemically defined fed-batch processes that support rapid cell proliferation, high cell density, and high production rate has been challenging.…”

Section: Introductionmentioning

confidence: 99%

A single nutrient feed supports both chemically defined NS0 and CHO fed‐batch processes: Improved productivity and lactate metabolism

Ellet

Okediadi

et al. 2009

Biotechnology Progress

112

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A chemically defined nutrient feed (CDF) coupled with basal medium preloading was developed to replace a hydrolysate-containing feed (HCF) for a fed-batch NS0 process. The CDF not only enabled a completely chemically defined process but also increased recombinant monoclonal antibody titer by 115%. Subsequent tests of CDF in a CHO process indicated that it could also replace the hydrolysate-containing nutrient feed in this expression system as well as providing an 80% increase in product titer. In both CDF NS0 and CHO processes, the peak lactate concentrations were lower and, more interestingly, lactate metabolism shifted markedly from net production to net consumption when cells transitioned from exponential to stationary growth phase. Subsequent investigations of the lactate metabolic shift in the CHO CDF process were carried out to identify the cause(s) of the metabolic shift. These investigations revealed several metabolic features of the CHO cell line that we studied. First, glucose consumption and lactate consumption are strictly complementary to each other. The combined cell specific glucose and lactate consumption rate was a constant across exponential and stationary growth phases. Second, Lactate dehydrogenase (LDH) activity fluctuated during the fed-batch process. LDH activity was at the lowest when lactate concentration started to decrease. Third, a steep cross plasma membrane glucose gradient exists. Intracellular glucose concentration was more than two orders of magnitude lower than that in the medium. Fourth, a large quantity of citrate was diverted out of mitochondria to the medium, suggesting a partially truncated tricarboxylic acid (TCA) cycle in CHO cells. Finally, other intermediates in or linked to the glycolytic pathway and the TCA cycle, which include alanine, citrate, isocitrate, and succinate, demonstrated a metabolic shift similar to that of lactate. Interestingly, all these metabolites are either in or linked to the pathway downstream of pyruvate, but upstream of fumarate in glucose metabolism. Although the specific mechanisms for the metabolic shift of lactate and other metabolites remain to be elucidated, the increased understanding of the metabolism of CHO cultures could lead to future improvements in medium and process development.

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“…However, the serum-free medium formulation used in this study was not published. In addition, many formulations for recombinant CHO cells expressing a variety of proteins are described (Asakura et al, 1992;Blüml et al, 1994;Hata et al, 1992;Ii et al, 1995;Kim et al, 1998;Kitano, 1994;Lee et al, 1999;Matthews et al, 1994;Mizuguchi et al, 1993;Scharfenberg and Wagner, 1995;Schorn et al, 1994;Wyatt, 1994). However, a versatile serum-free medium formulation for the parental cell line DUKXB11 has not been reported to date.…”

Section: Introductionmentioning

confidence: 99%

Serum- and protein-free media formulations for the Chinese hamster ovary cell line DUKXB11

Schröder

Matischak

Friedl

2004

Journal of Biotechnology

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Citation for published item:hr¤ oderD wF nd w tis h kD uF nd priedlD F @PHHRA 9 erumE nd proteinEfree medi formul tions for the ghinese h mster ov ry ell line h u fIIF9D tourn l of iote hnologyFD IHV @QAF ppF PUWEPWPF Further information on publisher's website: httpXGGdxFdoiForgGIHFIHITGjFj iote FPHHQFIPFHHS Publisher's copyright statement:Additional information: Use policyThe full-text may be used and/or reproduced, and given to third parties in any format or medium, without prior permission or charge, for personal research or study, educational, or not-for-pro t purposes provided that:• a full bibliographic reference is made to the original source • a link is made to the metadata record in DRO • the full-text is not changed in any way The full-text must not be sold in any format or medium without the formal permission of the copyright holders.Please consult the full DRO policy for further details. AbstractThe production of therapeutic proteins in mammalian cell lines is of outstanding importance. The maintenance of most mammalian cell lines in culture requires the addition of serum to the culture medium.The elimination of serum from mammalian cell culture is desirable since serum is expensive and a source of contaminants, e.g. viruses, mycoplasma or prions. Here we describe the composition of serum-and protein-free media for the Chinese hamster ovary (CHO) cell line DUKXB11. The serum-free formulation supports excellent growth of CHO DUKXB11 cells at low (23 cells/cm 2 ) and high (2·10 4 cells/cm 2 ) seeding densities characterized by a generation time of 10 -12 h, and, after addition of 0.2% pluronic F-68, the growth of a recombinant suspension cell line derived from DUKXB11. In addition, this formulation also allowed us to adapt recombinant cell lines expressing various amounts of human antithrombin ATIII (ATIII)to serum-free conditions. Secretion of ATIII was readily observed in the serum-free medium. Minor changes to the serum-free formulation resulted in a protein free formulation that supported growth of CHO DUKXB11 cells, growth of recombinant CHO cells expressing ATIII, and production of ATIII. Key wordsChinese hamster ovary cells, serum-free medium, protein-free medium, recombinant protein production,

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Chemically defined medium for the production of biologically active substances of CHO cells

Cited by 11 publications

References 14 publications

Fish glucocorticoid receptor with splicing variants in the DNA binding domain

Fish glucocorticoid receptor with splicing variants in the DNA binding domain

A single nutrient feed supports both chemically defined NS0 and CHO fed‐batch processes: Improved productivity and lactate metabolism

Serum- and protein-free media formulations for the Chinese hamster ovary cell line DUKXB11

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