Cell culture media for recombinant protein expression in Chinese hamster ovary (CHO) cells: History, key components, and optimization strategies

2018 ◽  
Vol 34 (6) ◽  
pp. 1407-1426 ◽  
Author(s):  
Frank V. Ritacco ◽  
Yongqi Wu ◽  
Anurag Khetan
Keyword(s):  
Cell Culture ◽  
Recombinant Protein ◽  
Protein Expression ◽  
Chinese Hamster Ovary ◽  
Cho Cells ◽  
Culture Media ◽  
Recombinant Protein Expression ◽  
Chinese Hamster ◽  
Cell Culture Media

scholarly journals Optimization of human interferon beta protein expression in Chinese hamster ovary cells

2021 ◽  
Vol 23 (2) ◽  
pp. 68-75
Author(s):  
Nasrin Xodadadi ◽  
Alireza Saeidinia ◽  
Mehdi Zeinoddini ◽  
Rasoul Khalilzadeh
Keyword(s):  
Recombinant Protein ◽  
Protein Expression ◽  
Chinese Hamster Ovary ◽  
Cho Cells ◽  
Interferon Beta ◽  
Recombinant Dna ◽  
Chinese Hamster Ovary Cells ◽  
Active Form ◽  
Chinese Hamster ◽  
Human Interferon

Background and aims: Human interferon beta-1a (hIFNβ-1a) is a 22.5-kDa glycoprotein used to treat diseases such as multiple sclerosis (MS). Because of appropriate post-translation modifications, protein isolation, and lack of toxicity in Chinese hamster ovary (CHO) cells, we cloned hIFNβ-1a encoding sequence into these cells by recombinant DNA technology to achieve stable expression of this recombinant protein. Methods: The hIFNβ-1a encoding sequence was designed based on the CHO cells’ codon usage and the Gene Bank data, and then syntactically constructed in the pUC57 vector. After confirmation, the synthesized sequence was cloned into the pcDNA3.1 expression vector by using EcoRI and XhoI sites via Escherichia coli DH5α competent cells. Then, the recombinant vector pcDNA-hHIFNβ1a was linearized by BglII and transfected into the CHO cells using lipofectamine. The transfected cells were proliferated and screened by gentamicin. Certain concentrations of zinc sulfate, DMSO, and glycerol were used to enhance protein expression. Finally, the recombinant protein expression was qualitatively evaluated using different techniques. Results: The hIFNβ1a integrity was confirmed by DNA sequencing and specific software. The construction and sub-cloning of hIFNβ1a-pcDNA3.1 in E. coli were confirmed by colony-PCR with specific primers and restriction enzyme mapping. The screening of transfected CHO cells was performed using gentamicin. The protein expression was confirmed by RT-PCR, MTT assay, SDS-PAGE, and Western blot. Comparison of the optimized and control samples demonstrated that chemical treatment enhanced the protein expression. Conclusion: We achieved the stable clones of CHO cells expressing the active form of human interferon beta.

scholarly journals A big cheese in biotherapeutics: Lactoyl leucine and isoleucine are bioavailable alternatives for canonical amino acids in cell culture media

2021 ◽  
Author(s):  
Corinna Schmidt ◽  
Maria Wehsling ◽  
Maxime Le Mignon ◽  
Gregor Wille ◽  
Yannick Rey ◽  
...  
Keyword(s):  
Amino Acids ◽  
Cell Culture ◽  
Cho Cells ◽  
Culture Media ◽  
Chinese Hamster ◽  
Batch Processes ◽  
Next Generation ◽  
Cell Culture Media ◽  
Derivatives Of

Increasing demands for protein-based therapeutics such as monoclonal antibodies, fusion proteins, bispecific molecules and antibody fragments require researchers to constantly find innovative solutions. To increase yields and decrease costs of next generation bioprocesses, highly concentrated cell culture media formulations are developed but often limited by the low solubility of amino acids such as tyrosine, cystine, leucine and isoleucine, in particular at physiological pH. This work sought to investigate highly soluble and bioavailable derivatives of leucine and isoleucine that are applicable for fed-batch processes. N-lactoyl-leucine and N-lactoyl-isoleucine sodium salts were tested in cell culture media and proved to be beneficial to increase the overall solubility of cell culture media formulations. These modified amino acids proved to be bioavailable for various Chinese hamster ovary (CHO) cells and were suitable for replacement of canonical amino acids in cell culture feeds. The quality of the final recombinant protein was studied in bioprocesses using the derivatives, and the mechanism of cleavage was investigated in CHO cells. Altogether, both N-lactoyl amino acids represent an advantageous alternative to canonical amino acids to develop highly concentrated cell culture media formulations to support next generation bioprocesses.

scholarly journals Advances of Glycometabolism Engineering in Chinese Hamster Ovary Cells

2021 ◽  
Vol 9 ◽  
Author(s):  
Huan-Yu Zhang ◽  
Zhen-Lin Fan ◽  
Tian-Yun Wang
Keyword(s):  
Protein Expression ◽  
Chinese Hamster Ovary ◽  
Cho Cells ◽  
Recombinant Protein Expression ◽  
Cell Metabolism ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Production Costs ◽  
Post Translational Modification ◽  
Ovary Cells

As the most widely used mammalian cell line, Chinese hamster ovary (CHO) cells can express various recombinant proteins with a post translational modification pattern similar to that of the proteins from human cells. During industrial production, cells need large amounts of ATP to support growth and protein expression, and since glycometabolism is the main source of ATP for cells, protein production partly depends on the efficiency of glycometabolism. And efficient glycometabolism allows less glucose uptake by cells, reducing production costs, and providing a better mammalian production platform for recombinant protein expression. In the present study, a series of progresses on the comprehensive optimization in CHO cells by glycometabolism strategy were reviewed, including carbohydrate intake, pyruvate metabolism and mitochondrial metabolism. We analyzed the effects of gene regulation in the upstream and downstream of the glucose metabolism pathway on cell’s growth and protein expression. And we also pointed out the latest metabolic studies that are potentially applicable on CHO cells. In the end, we elaborated the application of metabolic models in the study of CHO cell metabolism.

scholarly journals Comparison of the Production of Recombinant Protein in Suspension Culture of CHO Cells in Spinner Flask and Shake Flask System

1970 ◽  
Vol 12 (4) ◽  
Author(s):  
S.N.Z Zainul Abidin ◽  
And N. Anuar
Keyword(s):  
Recombinant Protein ◽  
Suspension Culture ◽  
Chinese Hamster Ovary ◽  
Cho Cells ◽  
Shake Flask ◽  
Specific Activity ◽  
Chinese Hamster ◽  
Lactate Production ◽  
Spinner Flask ◽  
Lac Z

Chinese hamster ovary (CHO) cells have been most widely used as the production host for the commercial production of biopharmaceuticals product. They have been extensively studied and developed, and today provide a stable platform for producing monoclonal antibodies and recombinant proteins. This study was focusing on comparison of suspension culture system by using spinner flask and shake flask for the growth and production of recombinant protein in CHO cell line. The CHO cells were transfected with an expression of DNA plasmid containing lac Z gene which codes for β-galactosidase. The recombinant genes in these CHO cells and the β-galactosidase expressing cells were adapted to suspension culture. The agitation speed for both spinner and shake flask were adjusted accordingly. The experiments were carried out in duplicate and samples were taken for cell count, determination of glucose consumption, lactate production and protein level by using biochemical assay. The result showed that, the cell growth in spinner flask is more favorable then in shake flask. The cell concentration in spinner flask is 58% higher than in shake flask. On the other hand, specific activity of β-galactosidase is 25% higher in spinner flask compared to shake flask, at the same agitation speed.ABSTRAK: Sel ovari hamster China (Chinese hamster ovary (CHO)) digunakan secara meluas dalam hos pembiakan untuk tujuan komersil produk biofarmaseutikal. Ia telah dikaji dan dibangunkan secara ekstensif, dan kini ia menyediakan landasan yang stabil untuk penghasilan antibodi monoklon dan protein rekombinan. Kajian ini memfokuskan tentang penghasilan protein rekombinan menggunakan kultur ampaian sel CHO di dalam kelalang putar dan kelalang goncang. Sel CHO dimasukkan dengan plasmid DNA yang mengandungi gen lac Z yang juga memberikan kod untuk β-galaktosidase. Sel CHO β-galaktosidase-terungkap dimasukkan ke dalam kultur ampaian. Kelajuan agitasi untuk kedua-dua kelalang putar dan kelalang goncang disesuaikan dengan sewajarnya. Eksperimen dijalankan menggunakan pendua dan sampel yang diambil untuk kiraan sel, penentuan penggunaan glukosa, penghasilan laktat dan aras protein dengan menggunakan cerakin biokimia. Keputusan menunjukkan tumbesaran sel di dalam kelalang putar lebih menggalakkan daripada dalam kelalang goncang. Kepekatan sel dalam kelalang putar adalah 58% lebih tinggi daripada dalam kelalang goncang. Sebaliknya, pada kelajuan agitasi yang sama, aktiviti tertentu β-galaktosidase adalah 25% lebih tinggi dalam kelalang putar dibandingkan dengan kelalang goncang.

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