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Culture medium used for Chinese hamster ovary cells

An ovarian cell, Chinese hamster technology, applied in the field of cell culture, can solve the problems of medium cost, low protein yield, high cost, etc.

Active Publication Date: 2012-09-05
SHANGHAI CELGEN BIO PHARMA CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Invitrogen also produces serum-free medium, but large-scale protein expression of antibody drugs requires a large amount of medium, which is expensive. For example, the price of JRH basic medium is more than 100 yuan per liter, which accounts for a large part of the cost. ratio, in my country, protein drug manufacturers are also faced with the problem of huge cost of culture medium, and at the same time, low protein production is also a big problem

Method used

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  • Culture medium used for Chinese hamster ovary cells
  • Culture medium used for Chinese hamster ovary cells
  • Culture medium used for Chinese hamster ovary cells

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0109] Preparation of basal medium for CHO cells

[0110] 1. Preparation of basal medium:

[0111] 1. Ingredients

[0112] Saijin basic CHO medium (prepared according to Table 1)

[0113] NaHCO 3 1.6g / L

[0114] L-Glutamine 0.4344g / L

[0115] Glucose 6.4g / L

[0116] Insulin (10mg / ml) 50ul ie: 0.5mg / L

[0117] 2. Filter Preparation

[0118] Ultra-pure water: the terminal resistance value of the water generator) is above 17 megohms, confirm that the balance is working normally, and the balance bubbles have been adjusted.

[0119] Select the filter according to the volume of the medium, and the suitable filter material: cartridge type hydrophilic filter element or flat filter membrane.

[0120]Cartridge filter element: 0.22um, the size is suitable for the sleeve, for integrity testing

[0121] Flat filter membrane: 0.45um, 0.22um each, fully wet

[0122] Filter sterilization: 122°C, 30 minutes

[0123] 3. Preparation

[0124] Add Saijin basic CHO medium, glucose an...

Embodiment 2

[0131] Formulating feed medium for CHO cells

[0132] 1. Preparation of fed-batch medium:

[0133] 1. Ingredients

[0134] Saijin feed medium (prepared according to the ratio in Table 2)

[0135] Glucose 24.4g / L

[0136] 2. Filter Preparation

[0137] Ultra-pure water: the terminal resistance value of the water generator) is above 17 megohms, confirm that the balance is working normally, and the balance bubbles have been adjusted.

[0138] Select the filter according to the volume of the medium, and the suitable filter material: cartridge type hydrophilic filter element or flat filter membrane.

[0139] Cartridge filter element: 0.22um, the size is suitable for the sleeve, for integrity testing

[0140] Flat filter membrane: 0.45um, 0.22um each, fully wet

[0141] Filter sterilization: 122°C, 30 minutes

[0142] 3. Preparation

[0143] Add Saijin feeding medium and glucose with 90% final volume of ultrapure water to dissolve, stir thoroughly for half an hour, dilute to...

Embodiment 3

[0149] Basal medium for culturing CHO cells

[0150] Test method: batch culture (without feeding)

[0151] Test procedure: In this culture verification, feeding is not carried out, and after inoculation, the cell survival rate decreases to about 50% as the end point of culture.

[0152] Culture environment: 37°C, 5% carbon dioxide

[0153] Shake flask culture process: Add the filtered basal medium and JRH medium of Example 1 to two 125ml shake flasks in a clean bench. Insert the seeds (counted as the 0th day), so that the final density of the cells is 0.5×10 6 , total volume 30ml. After the cells enter the middle stage of the plateau, the temperature begins to drop, so that the cells change from the growth state to the protein expression state.

[0154] Culture results: On the seventh day of culture with Saijin medium, the cell density reached the highest: 5.3x10 6 , the survival rate was 99.1%. While the cell density cultured with JRH basal medium was 3.9x10 6 , the su...

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Abstract

The invention discloses a basal culture medium used for Chinese hamster ovary cells. Based on the total volume of the basal culture medium, the basal culture medium comprises the following basic ingredients: 6-6.5g / L of sodium chloride, 0.5-0.5g / L of potassium chloride, 0.462-1g / L of pyruvic acid sodium, 0.288-0.4g / L of calcium nitrate, 1.464-2g / L of sodium bicarbonate, 5.16-15g / L of D-glucose and 0.156-0.7g / L of disodium hydrogen phosphate. The invention also discloses a fed-batch medium used for Chinese hamster ovary cells and a cultural method of Chinese hamster ovary cells.

Description

technical field [0001] The present invention relates to the field of cell culture. More specifically, the present invention relates to a medium for Chinese Hamster Ovary (CHO) cell culture and a corresponding culture method. Background technique [0002] CHO (Chinese Hamster Ovary) cells in eukaryotic cells are currently the preferred system for recombinant glycosyl protein production; because it has many advantages compared with other expression systems. At present, more and more medicinal proteins have been highly expressed in CHO cells, and many of the drugs expressed by CHO have been put on the market, such as EPO, Enbrel, etc. [0003] Currently, serum-free medium is widely used in CHO cell culture, because the medium containing animal serum has many disadvantages. Since the serum-free medium can only promote the growth and expression of cells, feed technology has now become one of the key points of competition among major biopharmaceutical companies. [0004] The co...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/071
Inventor 陈智胜
Owner SHANGHAI CELGEN BIO PHARMA CO LTD
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