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Low serum medium for full-suspending culture of MDCK cells

A medium and full suspension technology, applied in the direction of animal cells, culture process, tissue culture, etc., can solve the problems of low density and no medium, and achieve the effect of increasing cell density, low price, and increasing cell density

Inactive Publication Date: 2016-05-11
令世鑫 +3
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

According to reports, there have been many domesticated MDCK cells that can be fully cultured in suspension in my country, but the density of batch culture is not high, mostly at 2.0×10 6 About / mL, the main reason is that there is no corresponding complete suspension culture medium. In practical applications, many users buy serum-free culture at a high price and add 3-5% newborn bovine serum to normal culture cells.

Method used

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  • Low serum medium for full-suspending culture of MDCK cells
  • Low serum medium for full-suspending culture of MDCK cells
  • Low serum medium for full-suspending culture of MDCK cells

Examples

Experimental program
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Embodiment 1

[0049] The formula of the low-serum medium of the whole suspension culture MDCK cell of the present invention is as follows, the default unit: mg / L; wherein, the underlined part is the DMEM / F12 medium composition:

[0050] 1) Inorganic salts and trace elements:

[0051] Calcium Chloride CaCl 2 116.6

[0052] Potassium chloride KCl311.8

[0053] Magnesium Sulfate MgSO 4 48.84

[0054] Anhydrous disodium hydrogen phosphate Na 2 HPO 4 71.02

[0055] Sodium dihydrogen phosphate NaH 2 PO 4 -H 2 O62.5

[0056] Magnesium Chloride MgCl 2 28.64

[0057] Copper Sulfate CuSO 4 ·5H 2 O0.0013

[0058] Iron nitrate Fe(NO 3 ) 3 ·9H 2 O0.05

[0059] Ferrous Sulfate FeSO 4 ·7H 2 O0.417

[0060] Zinc sulfate ZnSO 4 ·7H 2 O0.432

[0061] Sodium pyruvate C 3 H 3 NaO 3 55

[0062] Nickel chloride NiCl 2 ·6H 2 O0.000195

[0063] Ammonium heptamolybdate (NH 4 ) 6 Mo 7 o 24 .4H 2 ...

Embodiment 2

[0123] The difference between this embodiment and embodiment 1 is:

[0124] In culture medium:

[0125] Sodium selenite 0.004mg / L

[0126] Sodium chloride NaCl6500mg / L

[0127] Vitamin A0.1mg / L

[0128]Vitamin E0.1mg / L

[0129] Human transferrin 5mg / L

[0130] Recombinant insulin 20mg / L

[0131] Glutathione 0.3mg / L

[0132] Vegetable protein hydrolyzate 2250mg / L

[0133] Recombinant human serum albumin 200mg / L

[0134] PF68800mg / L.

[0135] The remaining components of the culture medium and the preparation method of the culture medium are the same as in Example 1.

Embodiment 3

[0137] The difference between this embodiment and embodiment 1 is:

[0138] In culture medium:

[0139] Sodium selenite 0.003mg / L

[0140] Sodium chloride NaCl5000mg / L

[0141] Vitamin A0.2mg / L

[0142] Vitamin E0.08mg / L

[0143] Human transferrin 10mg / L

[0144] Recombinant insulin 15mg / L

[0145] Glutathione 0.1mg / L

[0146] Vegetable protein hydrolyzate 2500mg / L

[0147] Recombinant human serum albumin 300mg / L

[0148] PF681200mg / L.

[0149] The remaining components of the culture medium and the preparation method of the culture medium are the same as in Example 1.

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Abstract

The invention provides a low serum medium for full-suspending culture of MDCK cells. The medium is prepared from 5,000 mg / L to 6,500 mg / L of DMEM / F12 medium, nickel chloride NiCl2.6H2O, ammonium heptamolybdate (NH4) 6Mo7O24.4H2O, stannous chloride SnCl2.2H2O, ammonium metavanadate NH4VO3, sodium silicate Na2SiO3.9H2O, vitamin A, vitamin E, human transferrin, recombulin, glutathione, sodium selenite, vegetable protein hydrolysate, recombinant human serum albumin and PF68. New-born calf serum with the volume percent fraction of 3% to 5% is added to the medium, the medium is inoculated with the MDCK cells, cell density can be increased to 421*106 / ml, and the cell activity is still 92.3% at this moment.

Description

technical field [0001] The invention belongs to the technical field of biological products, and in particular relates to a low-serum culture medium for full suspension culture of MDCK cells. Background technique [0002] In recent years, large-scale outbreaks of influenza (avian flu) have seriously affected the health of humans and poultry animals. Vaccination is still the most important means of preventing influenza (avian flu). The production of traditional influenza vaccines uses the chicken embryo culture method, which has high probability of microbial contamination during the cultivation process, high endotoxin content of the product, large injection side effects, high cost of environmental pollution treatment of waste, difficulty in supplying chicken embryos when the influenza pandemic breaks out, and Due to the long production cycle and other technical shortcomings, it is imperative to develop an animal cell matrix vaccine for influenza. [0003] MDCK cells (Madin-Da...

Claims

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Application Information

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IPC IPC(8): C12N5/071
CPCC12N5/0625C12N2500/05C12N2500/24C12N2500/30C12N2500/32C12N2500/38C12N2500/84C12N2501/998C12N2501/999
Inventor 令世鑫马祺
Owner 令世鑫
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