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Cell culture method

A cell culture and culture medium technology, applied in the field of cell culture, can solve the problems of too much supernatant, flushing out of cells and carriers, limited amplification effect, etc.

Active Publication Date: 2013-09-18
SHANGHAI RITAI MEDICINE EQUIP PROJECT
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The present invention provides a cell culture method aiming at the defect that cells and carriers are easily flushed out by using the existing fluidized bed bioreactor for cell culture, and there are many remaining cells and carriers in the extracted supernatant, and the amplification effect is limited.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] Embodiment 1: Utilize the method of the present invention to carry out the mass cultivation of Vero cells

[0025] In the fluidized bed bioreactor (Shanghai Ritai 10L type), add sheet polyester carrier, add phosphate buffer (pH7.2) at the same time, sterilize under high pressure at 123 ℃ for 60 minutes, and naturally cool overnight. On the second day, the phosphate buffer solution in the tank was drained, and the DMEM medium containing 10% fetal bovine serum was added to inoculate Vero cells. After inoculation, the cell density in the tank reached 5x10 5 / ml to start cell culture.

[0026] The cell culture conditions of the reactor were set as temperature 37°C, pH 7.2, DO 50% for cell culture, the fluid passed through the fluidized bed from bottom to top, and flowed downward through the draft tube to form a swirling flow. Start the stirring device and set the rotation speed to 80 rpm. At this rotation speed, the fluid velocity is lower than the flushing point, forming ...

Embodiment 2

[0030] Embodiment 2: Utilize the method of the present invention to carry out the mass culture of Chinese hamster ovary cell (CHO cell)

[0031] Add 300 grams of domestic flake polyester carrier and 8 L of phosphate buffer (pH 7.2) into a 10 L fluidized bed bioreactor, autoclave at 123 °C for 60 minutes, and cool overnight naturally. On the second day, the phosphate buffer solution in the tank was drained, and DMEM medium containing 10% fetal bovine serum was added to inoculate CHO cells. After inoculation, the cell density in the tank reached 5x10 5 / ml to start cell culture.

[0032] The cell culture conditions of the reactor were set as temperature 37°C, pH 7.2, DO 50% for cell culture, the fluid passed through the fluidized bed from bottom to top, and flowed downward through the draft tube to form a swirling flow. Start the stirring device and set the rotation speed to 80 rpm. At this rotation speed, the fluid velocity is lower than the flushing point, forming a culture m...

Embodiment 3

[0036] Embodiment 3: Utilize the method of the present invention to carry out the mass culture of 293 cells

[0037] Add 300 grams of domestic flake polyester carrier and 8 L of phosphate buffer (pH 7.2) into a 10 L fluidized bed bioreactor, autoclave at 123 °C for 60 minutes, and cool overnight naturally. On the second day, the phosphate buffer solution in the tank was drained, and high-sugar DMEM medium was added to inoculate 293 cells. After inoculation, the cell density in the tank reached 5x10 5 / ml to start cell culture.

[0038] The cell culture conditions of the reactor were set as temperature 37°C, pH 7.2, DO 50% for cell culture, the fluid passed through the fluidized bed from bottom to top, and flowed downward through the draft tube to form a swirling flow. Start the stirring device and set the rotation speed to 80 rpm. At this rotation speed, the fluid velocity is lower than the flushing point, forming a culture method similar to a fixed bed, but a slight shaking ...

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PUM

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Abstract

The invention relates to the technical field of biology, and discloses a cell culture method. The cell culture method disclosed by the invention can homogenize nutrient substances and metabolites of cells in the tank body of a fluidized bed bioreactor, so that the cell density can reach higher than 10<7> / ml, thereby implementing continuous culture. The cell culture method disclosed by the invention is free from the restrictions of conditions, and has great scale-up potential and wide industrial application prospects.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a cell culture method. Background technique [0002] Currently, cell culture is commonly carried out in fluidized bed bioreactors or fixed bed bioreactors. The fixed bed bioreactor is a fixed container installed in the reactor, and the carrier is filled in the middle. The carrier is usually a solid or porous ball or sheet-like fiber with a diameter of 2-6mm. The culture medium is circulated through the fixed bed, relying on the negative pressure generated during the stirring, so that the medium continuously flows through the filler for the transfer of nutrients and oxygen. The freshly inoculated cells grow on the surface of the carrier, and as the cells proliferate, the cells begin to fill the spaces between the carriers. [0003] The fixed-bed bioreactor culture perfusion speed is relatively high, and the cells grow at a high density in the carrier, which can reduce the amount of ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/071
Inventor 郑振彬陈光南
Owner SHANGHAI RITAI MEDICINE EQUIP PROJECT
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