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Animal cell feeding culture process

A fed-batch culture, animal cell technology, applied in animal cells, fusion cells, cells modified by introducing foreign genetic material, etc., can solve the problems of divergence, long research cycle, process instability and so on

Inactive Publication Date: 2006-05-17
EAST CHINA UNIV OF SCI & TECH
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Apparently, although the method of Xie et al. seems to be accurate, it is too complicated and will inevitably lead to a long research cycle and a huge workload.
At the same time, in the fed-batch culture process of animal cells, the metabolic rules and physiological state of the cells are not constant, so neither Xie nor Zhou's method design specifically considers the feedback and correction of the process. Once the difference exists, it will be Cause process instability, even divergence, leading to eventual failure

Method used

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Experimental program
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Effect test

Embodiment 1～2

[0084] (a). Metabolism study

[0085] The physiological state of HB58 hybridoma cells (ATCC) under glucose-limited or glutamine-limited culture conditions is studied by using the method described in the present invention. Inoculate in Biostat B (Germany B.BRAUN company) 2 liters of bioreactors, inoculation density 2.0 * 10 5 cells / ml. When the living cell density reaches 1.0×10 6 Start fed-batch culture after cells / ml, the dilution rate is 0.30day -1 , the culture time was 125 hours and 100 hours respectively, achieving glucose limitation (see figure 2 and image 3 ) and glutamine-limited quasi-steady-state culture ( Figure 4 and Figure 5 ), can be used to study the metabolic rules when the cells are in the corresponding state. During the whole cultivation process, the stirring speed of the reactor was 120 rpm, the temperature was 36.8° C., and the dissolved oxygen was 50% air saturation.

[0086] Element

Inorganic salt (mg / L)

CaCl 2 ...

Embodiment 3～4

[0093] (b). High-density optimized fed-batch culture

[0094] HB58 hybridoma cells (ATCC) were optimized fed-batch culture by the method of the present invention. In a Biostat B (Germany B.BRAUN company) 2 liter bioreactor, the live cell seeding density was 2.0×10 5cells / ml. The concentrations of glucose and glutamine in the initial medium were 7.8mmol / L and 2.5mmol / L, respectively, and the concentrations of other amino acids were the original concentrations of DMEM / F12 (1:1) medium. When the viable cell density reaches 1.0×10 6 After cells / ml, start feeding medium exponentially based on cell demand. The feeding dilution rate is 0.30day -1 .

[0095] The concentrations of glucose and glutamine in fed-batch medium were 100mmol / L and 33mmol / L, respectively, and the amino acid was 6.5 times the mass of DMEM / F12 (1:1, mass ratio) medium. During the whole cultivation process, the stirring speed of the reactor was 120 rpm, the temperature was 36.8° C., and the dissolved oxygen...

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Abstract

The animal cell feeding culture process includes the following steps: designing different feeding processes to realize different pseudostable culture environment, determine optimized target and establish metabolism dynamics model; performing the optimized culture process based on the metabolism dynamics model; sampling and detecting ammonia concentration regularly, estimating the concentration of glutamine as the control quantity based on the metabolism dynamics model and feeding back to correct feeding rate to avoid system divergency. The process of the present invention can perform research on metabolism regulations of cell strain fast and accurately and realize cell culture process with high culture medium efficiency, high cell density and product concentration, high stability and controllability. The present invention makes it possible to establish large scale optimized animal cell feeding culture process and is significant in producing bioproduct.

Description

technical field [0001] The invention relates to a method for large-scale, high-density fed-batch culture of animal cells. technical background [0002] Large-scale culture of animal cells has been widely used in the production of various biologically active substances such as monoclonal antibodies, virus vaccines, immune regulators, growth factors, specific tumor antigens, and various gene recombinant protein drugs. Compared with microorganisms, animal cells have the ability of post-transcriptional modification, and can efficiently express and produce various high-quality proteins. However, the medium used for animal cell culture is expensive, and the low expression level results in high cost of separation and purification. Therefore, increasing cell density and product concentration and reducing medium consumption are of great significance in industrial applications. [0003] In simple batch culture (Batch), with the consumption of nutrients and the accumulation of by-pro...

Claims

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Application Information

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IPC IPC(8): C12N5/07C12N5/12C12N5/16
Inventor 谭文松牛红星朱明龙周燕华平顾小华
Owner EAST CHINA UNIV OF SCI & TECH
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