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Method for harvesting cell culture fluid

A technology of cell culture and harvesting method, applied in the field of cell culture fluid harvesting, which can solve problems such as production failure, increased production equipment and facility operating load, and reduced product market competitiveness

Pending Publication Date: 2021-06-04
INNOVENT BIOLOGICS (SUZHOU) CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] In the prior art, the ability to increase the capacity of depth filtration membrane packs is limited, and specific membrane packs need to be matched, and the number of membrane packs used is significantly larger than that of similar products, thereby increasing the operating load of production equipment and facilities, and there is a possibility of production failure risk
In addition, the use of deep filter membranes to harvest cell culture fluid will inevitably increase the production cost of the product and reduce the market competitiveness of the product.

Method used

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  • Method for harvesting cell culture fluid
  • Method for harvesting cell culture fluid
  • Method for harvesting cell culture fluid

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0079] Embodiment 1: cell culture basic technique

[0080] This example describes the cell culture process applicable to the present invention. In this example, the target protein in the cell harvesting solution is a recombinant fully human anti-programmed death receptor 1 (PD-1) bispecific antibody.

[0081] The basic process of cell culture process includes:

[0082] 1) Seed recovery: Take a working cell expressing anti-PD-1 bispecific antibody from the working cell bank, thaw it in a water bath at 37°C±0.5°C, transfer the cell suspension to a centrifuge tube, add 10mL Centrifuge the basal medium M at 1000r / min for 5 minutes, remove the supernatant, resuspend the cells with basal medium M and transfer to a 125mL flat-bottomed shaker flask, replenish the volume to 30mL, mix well and take a sample for cell counting. Place the shaker flask in a carbon dioxide shaker and incubate at 36.5°C ± 0.5°C, 6% ± 1% CO 2 , under the condition of 130±10r / min, cultivate for 3±1 days.

[...

Embodiment 2

[0086] Example 2: The performance of the cell culture solution containing the bispecific antibody against the PD-1 receptor in the depth filtration process stage under the condition that the traditional two-stage depth filtration membrane package is connected in series

[0087] A) Assembly of depth filtration membrane: Membrane bag D0HC (Millipore company product, article number MDOHC23CL3, membrane dead volume 23mL), membrane bag X0HC (Millipore company product, article number: MXOHC23CL3, membrane dead volume 23mL), use silicone tube (NALGENE Company product, article number: 8600-0030), successively connected in series peristaltic pump (product of Intertek company, model: 120s)-pressure gauge (product of Millipore company, article number: SLTEST000)-D0HC membrane bag-X0HC membrane bag, pumped into the buffer by the peristaltic pump solution or sample, and finally flows out from the XOHC membrane capsule.

[0088]B) Rinse the depth filtration membrane bag with ultrapure water...

Embodiment 3

[0102] Example 3: The performance of cell culture fluid containing bispecific antibody against PD-1 receptor in the depth filtration process stage under the condition of simulated continuous flow centrifuge centrifugation

[0103] A) Assembly of depth filtration membrane: Membrane bag D0HC (product of millipore company, article number MDOHC23CL3, membrane dead volume 23mL), membrane bag X0HC (millipore company product, article number: MXOHC23CL3, membrane dead volume 23mL), use silicone tube (NALGENE Company product, article number: 8600-0030) in series with peristaltic pump (Intertek company product, model: 120s)-pressure gauge (Millipore company product, article number: SLTEST000)-D0HC membrane bag-X0HC membrane bag, pumped into the buffer by the peristaltic pump or samples, and finally flow out from the XOHC membrane capsule.

[0104] B) Centrifugation: The centrifugal force parameter is set to 2000G centrifugal force and centrifuged for 3 minutes; the cell density obtained...

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Abstract

The invention discloses a method for harvesting a cell culture fluid. The method comprises the following steps of 1) providing host cells, and carrying out seed recovery, shake-flask multiplication culture and fed-batch culture on the host cells to obtain a cell culture with the cell density of not less than 1.0*10<6> cells / mL; 2) adding an acidic solution into the cell culture in the step 1), and adjusting the pH value of the cell culture to 4.0 to 5.6; 3) deeply filtering the cell culture in the step 2) and / or adding an alkaline solution to obtain the cell culture fluid; and 4) harvesting the cell culture fluid in the step 3). By utilizing the method for harvesting the cell culture fluid provided by the invention, the harvesting efficiency of the cell culture fluid can be remarkably improved, meanwhile, the entrapment capacity of a deep filtering membrane can be remarkably improved, the end-point turbidity of the cell culture fluid is reduced, the operability of production is improved, and the cost investment of production equipment and facilities is reduced.

Description

technical field [0001] The invention relates to the field of cell culture, in particular to a method for harvesting cell culture fluid. Background technique [0002] After decades of development, cell culture technology has become a mature technology for the production of many important proteins. Currently, commonly used cell culture fluid harvesting methods in the field of cell culture include: flocculation, sedimentation, centrifugation and depth filtration. Among them, centrifugation and deep layer filtration cannot effectively separate the cell harvest liquid of some specific medium components, and the cell harvest liquid obtained by centrifugation or deep layer filtration has high turbidity. In addition, the low loading capacity of the depth filtration membrane used in the depth filtration method will also affect the harvest of the cell culture medium and the yield of the target product. The cell culture medium is harvested by flocculation or sedimentation, and its ma...

Claims

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Application Information

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IPC IPC(8): C12N5/10C07K16/46
CPCC12N5/0682C07K16/2818C12N2500/34C12N2500/32C12N2510/02C07K2317/24C07K2317/31C07K2317/14
Inventor 杨忠华秦婷王亚东黄海峰王晓玉
Owner INNOVENT BIOLOGICS (SUZHOU) CO LTD
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