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CN101195816A - Gonad cell amplification culture medium of Chinese hamster and uses thereof - Google Patents

Gonad cell amplification culture medium of Chinese hamster and uses thereof Download PDF

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Publication number
CN101195816A
CN101195816A CNA2007103070769A CN200710307076A CN101195816A CN 101195816 A CN101195816 A CN 101195816A CN A2007103070769 A CNA2007103070769 A CN A2007103070769A CN 200710307076 A CN200710307076 A CN 200710307076A CN 101195816 A CN101195816 A CN 101195816A
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CN
China
Prior art keywords
water
composition
acid
sodium
vitamin
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CNA2007103070769A
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Chinese (zh)
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CN100506977C (en
Inventor
黄宗堂
唐贺文
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TIANJIN BAINUOKE MEDICINE BIOTECHNOLOGY CO Ltd
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TIANJIN BAINUOKE MEDICINE BIOTECHNOLOGY CO Ltd
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Priority to CNB2007103070769A priority Critical patent/CN100506977C/en
Publication of CN101195816A publication Critical patent/CN101195816A/en
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Abstract

The invention relates to a culture medium and the usage thereof, in particular to the Chinese hamster ovary cell enlarging culture medium and the usage thereof. The invention belongs to the biological product field. The Chinese hamster ovary cell enlarging non-protein culture medium of the invention, wherein comprises minimal medium, and also comprises microelement, Transferrin and Insulin subsistent. A serum-containing routine culture medium can be directly replaced with the serum free culture medium of the Chinese hamster ovary cells, and the Chinese hamster ovary cells are not required to adapt process.

Description

A kind of Chinese hamster ovary cell amplification culture medium and uses thereof
Technical field
Invention relates to a kind of substratum and uses thereof, more specifically to a kind of Chinese hamster ovary cell amplification culture medium and uses thereof.Belong to field of biological product.
Background technology
Cell culture medium is material commonly used in the field of biological product, and is very necessary in bacterium and cell cultures.The shortcoming of cultivating the substratum of Chinese hamster ovary cell at present is: at present, animal serum is widely used as the key component of mammalian cell substratum, based on bovine serum.But the animal serum shortcoming is: cost an arm and a leg, composition is indeterminate, and the product differences between batches are big, contains cell growth-inhibiting composition, and even more serious be possible contain bacterium, virus etc., cause the potential source biomolecule contact scar.
Summary of the invention
[problem that will solve]
Purpose of the present invention provides a kind of Chinese hamster ovary cell amplification culture medium and uses thereof.
[technical scheme]
A kind of Chinese hamster ovary cell of the present invention (Chinese hamster ovary celI) amplifying protein-free substratum wherein, comprises minimum medium, also comprises: trace element and Transferrin Transferrins,iron complexes and Insulin insulin substitution thing.
Above-mentioned minimum medium comprises: amino acid, VITAMIN, inorganic salt, other nutritive substances.
Component that above-mentioned minimum medium comprises and content are:
Amino acid whose composition and weight part are as follows:
Composition Mg/L
Glycine 2-30
L-L-Ala 3-40
L-arginine monohydrochloride 50-450
One water L-amino-succinamic acid 3.0-45.0
L-aspartic acid 5.0-2 5.0
L-cysteine hydrochloride 3.0-45.0
One water L-cysteine hydrochloride 10.0-65.0
L-L-glutamic acid 4-35
One water L-L-Histidine hydrochloride 10-75
L-Isoleucine 20-100
L-leucine 15-150
L lysine HCL 50-300
L-methionine(Met) 3-40
L-phenylalanine 1.0-15
L-proline(Pro) 15-85
L-Serine 3.0-30
L-Threonine 3.0-30
L-tryptophane 0.5-8
L-tyrosine disodium two water 3.0-25
L-Xie Ansuan 15-100
The composition and the weight part of VITAMIN are as follows:
Composition Mg/L
Vitamin H 0.0005-0.09
Choline chloride 60 3.0-20
D-calcium pantothenate 0.5-9
Folic acid 0.5-10
Inositol 3.0-35
Niacinamide 3.0-25
Pyridoxal hydrochloride 0.1-5
Pyridoxine hydrochloride 10-100
Riboflavin 0.1-3
The plain hydrochloride 0.1-3 of nuclear ammonia
Vitamins B 120.001-0.9
Vitamins C 0.001-0.2
Vitamin K 30.001-0.2
Nicotinic acid 0.008-0.3
Vitamin A acetic acid 0.009-0.3
Vitamins D 20.001-0.2
The composition and the weight part of inorganic salt are as follows:
Composition Mg/L
Calcium chloride 45-250
Magnesium chloride 10-65
Sal epsom 20-100
Repone K 100-800
Sodium-chlor 2500-10000
Sodium dihydrogen phosphate-water 25-150
Sodium phosphate dibasic 25-150
Sodium bicarbonate 900-5000
The composition and the weight part of other nutritive substances are as follows:
Composition Mg/L
Glucose 2000-6500
Phenol red 0.5-15
Sodium.alpha.-ketopropionate 60-300
Xanthine 0.5-4.5
Linolic acid 0.001-0.5
Thioctic Acid 0.005-0.8
Putrescine acid 0.001-0.25
Gsh 0.1-1.5
Thymidine 0.08-0.85
4-hydroxyethyl piperazine ethanesulfonic acid 800-3000
Thanomin 0.4-4.0
Preferred component and content that above-mentioned minimum medium comprises are:
Amino acid whose composition and weight part are as follows:
Composition Mg/L
Glycine 7.5
L-L-Ala 8.9
L-arginine monohydrochloride 211
One water L-amino-succinamic acid 15.01
L-aspartic acid 13.3
L-cysteine hydrochloride 17.56
One water L-cysteine hydrochloride 31.29
L-L-glutamic acid 7.35
One water L-L-Histidine hydrochloride 31.48
L-Isoleucine 54.47
L-leucine 59.05
L lysine HCL 91.25
L-methionine(Met) 17.24
L-phenylalanine 5
L-proline 3 4.5
L-Serine 10.5
L-Threonine 11.9
L-tryptophane 2.04
L-tyrosine disodium two water 7.81
L-Xie Ansuan 46
The composition and the weight part of VITAMIN are as follows:
Composition Mg/L
Vitamin H 0.0035
Choline chloride 60 8.98
D-calcium pantothenate 2.24
Folic acid 2.65
Inositol 18
Niacinamide 12
Pyridoxal hydrochloride 1
Pyridoxine hydrochloride 35
Riboflavin 1
The plain hydrochloride 1 of nuclear ammonia
Vitamin B12 0.01
Vitamins C 0.05
Vitamin K3 0.05
Nicotinic acid 0.025
Vitamin A acetic acid 0.025
Vitamin D2 0.01
The composition and the weight part of inorganic salt are as follows:
Composition Mg/L
Calcium chloride 116.6
Magnesium chloride 28.64
Sal epsom 48.84
Repone K 311.8
Sodium-chlor 6995.5
Sodium dihydrogen phosphate-water 62.5
Sodium phosphate dibasic 71.02
Sodium bicarbonate 2200
The composition and the weight part of other nutritive substances are as follows:
Composition Mg/L
Glucose 3900
Phenol red 4
Sodium.alpha.-ketopropionate 150
Xanthine 1.66
Linolic acid 0.035
Thioctic Acid 0.075
Putrescine acid 0.004
Gsh 0.6
Thymidine 0.28
4-hydroxyethyl piperazine ethanesulfonic acid 1800
Thanomin 1.2
Component that above-mentioned trace element comprises and content are:
Component Mg/L
Cupric sulfate pentahydrate 0.0009-0.02
Sodium Fluoride 0.0008-0.01
Nine water Sodium Selenite 0.05-0.2
Barium acetate 0.001-0.03
Eight water Cadmium Sulphate 0.0007-0.004
Zirconium oxychloride 0.001-0.0085
Component that above-mentioned trace element comprises and content are:
Component Mg/L
Cupric sulfate pentahydrate 0.00182
Sodium Fluoride 0.001 97
Nine water Sodium Selenites 0.094
Barium acetate 0.0079
Eight water Cadmium Sulphates 0.00122
Zirconium oxychloride 0.0015
Component and content that above-mentioned Transferrin Transferrins,iron complexes and Insulin Regular Insulin albumen surrogate comprise are:
Composition Mg/L
Zinc sulfate 1.0-15
1,2-arachidonic acyl Yelkin TTS 1.0-16
Ferrous sulfate seven water 0.05-0.12
Iron nitrate nine water water 0.01-0.45
Ironic oxalate 0.1-2.0
Ethylenediamine tetraacetic acid (EDTA) 4 sodium 0.01-0.45
Component and content that above-mentioned Transferrin Transferrins,iron complexes and Insulin Regular Insulin albumen surrogate comprise are:
Composition Mg/L
Zinc sulfate 5
1,2-arachidonic acyl Yelkin TTS 5
Ferrous sulfate seven water 0.094
Iron nitrate nine water water 0.05
Ironic oxalate 0.6
Ethylenediamine tetraacetic acid (EDTA) 4 sodium 0.03
Above-mentioned substratum has purposes in cultivating Chinese hamster ovary cell.
Above-mentioned substratum, this Chinese hamster ovary cell amplification serum free medium can directly be replaced and contains the conventional substratum of serum, and Chinese hamster ovary cell does not need adaptive process.
The advantage of product of the present invention is: Chinese hamster ovary cell protein-free medium of the present invention, do not contain any animal serum, and with amino acid, VITAMIN, inorganic salt, carbohydrate, lipid acid and trace element are formed.Do not contain Transferrins,iron complexes and Regular Insulin, these two kinds of compositions.With ZnSO 4, 1,2-Diarachidonoyl-Phosphatidylcholine, FeSO 47H 2O, Fe (NO 3) 39H 2O, Ferric Oxalate, EDTA 4Na replaces.But these alternative compositions are synthetic all, is the clear and definite chemical substance of structure, makes the basal culture medium product have high consistence and stability.Transferrins,iron complexes and Regular Insulin, no matter animal derived extraction purifying is originated or through the acquisition of recombinant protein technology synthetic, is all cost an arm and a leg, and after substituting with above component, culture medium cost significantly reduces.In addition,, make that its downstream purification work was greatly simplified when the Chinese hamster ovary cell of using the basal culture medium cultivation carried out recombinant protein production, save cost, be beneficial to the purity that improves recombinant protein because of basal culture medium does not contain protein component.
This Chinese hamster ovary cell serum free medium can directly be replaced and contains the conventional substratum of serum, and Chinese hamster ovary cell does not need adaptive process.
Description of drawings
Fig. 1 is the cultivation curve of the embodiment 3 of serum free medium Chinese hamster ovary cell amplification culture medium;
Fig. 2 is the cultivation curve of the embodiment 2 of serum free medium Chinese hamster ovary cell amplification culture medium.
Embodiment
Embodiment 1
Substratum is made up of minimum medium, trace element and Transferrin Transferrins,iron complexes and Insulin insulin substitution thing, and wherein, the composition and the composition of minimum medium are as follows:
Amino acid whose composition and weight part are as follows:
Composition Mg/L
Glycine 7.5
L-L-Ala 8.9
L-arginine monohydrochloride 211
One water L-amino-succinamic acid 15.01
L-aspartic acid 13.3
L-cysteine hydrochloride 17.56
One water L-cysteine hydrochloride 31.29
L-L-glutamic acid 7.35
One water L-L-Histidine hydrochloride 31.48
L-Isoleucine 54.47
L-leucine 59.05
L lysine HCL 91.25
L-methionine(Met) 17.24
L-phenylalanine 5
L-proline 3 4.5
L-Serine 10.5
L-Threonine 11.9
L-tryptophane 2.04
L-tyrosine disodium two water 7.81
L-Xie Ansuan 46
The composition and the weight part of VITAMIN are as follows:
Composition Mg/L
Vitamin H 0.0035
Choline chloride 60 8.98
D-calcium pantothenate 2.24
Folic acid 2.65
Inositol 18
Niacinamide 12
Pyridoxal hydrochloride 1
Pyridoxine hydrochloride 35
Riboflavin 1
The plain hydrochloride 1 of nuclear ammonia
Vitamin B12 0.01
Vitamins C 0.05
Vitamin K3 0.05
Nicotinic acid 0.025
Vitamin A acetic acid 0.025
Vitamin D2 0.01
The composition and the weight part of inorganic salt are as follows:
Composition Mg/L
Calcium chloride 116.6
Magnesium chloride 28.64
Sal epsom 48.84
Repone K 311.8
Sodium-chlor 6995.5
Sodium dihydrogen phosphate-water 62.5
Sodium phosphate dibasic 71.02
Sodium bicarbonate 2200
The composition and the weight part of other nutritive substances are as follows:
Composition Mg/L
Glucose 3900
Phenol red 4
Sodium.alpha.-ketopropionate 150
Xanthine 1.66
Linolic acid 0.035
Thioctic Acid 0.075
Putrescine acid 0.004
Gsh 0.6
Thymidine 0.28
4-hydroxyethyl piperazine ethanesulfonic acid 1800
Thanomin 1.2
Component that trace element comprises and content are:
Component Mg/L
Cupric sulfate pentahydrate 0.00182
Sodium Fluoride 0.00197
Nine water Sodium Selenites 0.094
Barium acetate 0.0079
Eight water Cadmium Sulphates 0.00122
Zirconium oxychloride 0.0015
Component and content that Transferrin Transferrins,iron complexes and Insulin Regular Insulin albumen surrogate comprise are:
Composition Mg/L
Zinc sulfate 5
1,2-arachidonic acyl Yelkin TTS 5
Ferrous sulfate seven water 0.094
Iron nitrate nine water water 0.05
Ironic oxalate 0.6
Ethylenediamine tetraacetic acid (EDTA) 4 sodium 0.03
Embodiment 2
Substratum is made up of minimum medium, trace element and Transferrin Transferrins,iron complexes and Insulin insulin substitution thing, and wherein, the composition and the composition of minimum medium are as follows:
Amino acid whose composition and weight part are as follows:
Composition Mg/L
Glycine 2
L-L-Ala 3
L-arginine monohydrochloride 50
One water L-amino-succinamic acid 3.0
L-aspartic acid 5.0
L-cysteine hydrochloride 3.0
One water L-cysteine hydrochloride 10.0
L-L-glutamic acid 4
One water L-L-Histidine hydrochloride 10
L-Isoleucine 20
L-leucine 15
L lysine HCL 50
L-methionine(Met) 3
L-phenylalanine 1.0
L-proline(Pro) 15
L-Serine 3.0
L-Threonine 3.0
L-tryptophane 0.5
L-tyrosine disodium two water 3.0
L-Xie Ansuan 15
The composition and the weight part of VITAMIN are as follows:
Composition Mg/L
Vitamin H 0.0005
Choline chloride 60 3.0
D-calcium pantothenate 0.5
Folic acid 0.5
Inositol 3.0
Niacinamide 3.0
Pyridoxal hydrochloride 0.1
Pyridoxine hydrochloride 10
Riboflavin 0.1
The plain hydrochloride 0.1 of nuclear ammonia
Vitamin B12 0.001
Vitamins C 0.001
Vitamin K3 0.001
Nicotinic acid 0.008
Vitamin A acetic acid 0.009
Vitamin D2 0.001
The composition and the weight part of inorganic salt are as follows:
Composition Mg/L
Calcium chloride 45
Magnesium chloride 10
Sal epsom 20
Repone K 100
Sodium-chlor 2500
Sodium dihydrogen phosphate-water 25
Sodium phosphate dibasic 25
Sodium bicarbonate 900
The composition and the weight part of other nutritive substances are as follows:
Composition Mg/L
Glucose 2000
Phenol red 0.5
Sodium.alpha.-ketopropionate 60
Xanthine 0.5
Linolic acid 0.001
Thioctic Acid 0.005
Putrescine acid 0.001
Gsh 0.1
Thymidine 0.08
4-hydroxyethyl piperazine ethanesulfonic acid 800
Thanomin 0.4.
Component that trace element comprises and content are:
Component Mg/L
Cupric sulfate pentahydrate 0.0009
Sodium Fluoride 0.0008
Nine water Sodium Selenites 0.05
Barium acetate 0.001
Eight water Cadmium Sulphates 0.0007
Zirconium oxychloride 0.001.
Component and content that Transferrin Transferrins,iron complexes and Insulin Regular Insulin albumen surrogate comprise are:
Composition Mg/L
Zinc sulfate 1.0
1,2-arachidonic acyl Yelkin TTS 1.0
Ferrous sulfate seven water 0.05
Iron nitrate nine water water 0.01
Ironic oxalate 0.1
Ethylenediamine tetraacetic acid (EDTA) 4 sodium 0.01.
Embodiment 3
Substratum is made up of minimum medium, trace element and Transferrin Transferrins,iron complexes and Insulin insulin substitution thing, and wherein, the composition and the composition of minimum medium are as follows:
Amino acid whose composition and weight part are as follows:
Composition Mg/L
Glycine 30
L-L-Ala 40
L-arginine monohydrochloride 450
One water L-amino-succinamic acid 45.0
L-aspartic acid 25.0
L-cysteine hydrochloride 45.0
One water L-cysteine hydrochloride 65.0
L-L-glutamic acid 35
One water L-L-Histidine hydrochloride 75
L-Isoleucine 100
L-leucine 150
L lysine HCL 300
L-methionine(Met) 40
L-phenylalanine 15
L-proline(Pro) 85
L-Serine 30
L-Threonine 30
L-tryptophane 8
L-tyrosine disodium two water 25
L-Xie Ansuan 100
The composition and the weight part of VITAMIN are as follows:
Composition Mg/L
Vitamin H 0.09
Choline chloride 60 20
D-calcium pantothenate 9
Folic acid 10
Inositol 35
Niacinamide 25
Pyridoxal hydrochloride 5
Pyridoxine hydrochloride 100
Riboflavin 3
The plain hydrochloride 3 of nuclear ammonia
Vitamin B12 0.9
Vitamins C 0.2
Vitamin K3 0.2
Nicotinic acid 0.3
Vitamin A acetic acid 0.3
Vitamin D2 0.2
The composition and the weight part of inorganic salt are as follows:
Composition Mg/L
Calcium chloride 250
Magnesium chloride 65
Sal epsom 100
Repone K 800
Sodium-chlor 10000
Sodium dihydrogen phosphate-water 150
Sodium phosphate dibasic 150
Sodium bicarbonate 5000
The composition and the weight part of other nutritive substances are as follows:
Composition Mg/L
Glucose 6500
Phenol red 15
Sodium.alpha.-ketopropionate 300
Xanthine 4.5
Linolic acid 0.5
Thioctic Acid 0.8
Putrescine acid 0.25
Gsh 1.5
Thymidine 0.85
4-hydroxyethyl piperazine ethanesulfonic acid 3000
Thanomin 4.0.
Component that trace element comprises and content are:
Component Mg/L
Cupric sulfate pentahydrate 0.02
Sodium Fluoride 0.01
Nine water Sodium Selenites 0.2
Barium acetate 0.03
Eight water Cadmium Sulphates 0.004
Zirconium oxychloride 0.0085.
Component and content that Transferrin Transferrins,iron complexes and Insulin Regular Insulin albumen surrogate comprise are:
Composition Mg/L
Zinc sulfate 15
1,2-arachidonic acyl Yelkin TTS 16
Ferrous sulfate seven water 0.12
Iron nitrate nine water water 0.45
Ironic oxalate 2.0
Ethylenediamine tetraacetic acid (EDTA) 4 sodium 0.45.
Embodiment 4
Substratum is made up of minimum medium, trace element and Transferrin Transferrins,iron complexes and Insulin insulin substitution thing, and wherein, the composition and the composition of minimum medium are as follows:
Amino acid whose composition and weight part are as follows:
Composition Mg/L
Glycine 2
L-L-Ala 40
L-arginine monohydrochloride 50
One water L-amino-succinamic acid 45.0
L-aspartic acid 5.0
L-cysteine hydrochloride 45.0
One water L-cysteine hydrochloride 10.0
L-L-glutamic acid 35
One water L-L-Histidine hydrochloride 10
L-Isoleucine 100
L-leucine 15
L lysine HCL 300
L-methionine(Met) 3
L-phenylalanine 15
L-proline(Pro) 15
L-Serine 30
L-Threonine 3.0
L-tryptophane 8
L-tyrosine disodium two water 3.0
L-Xie Ansuan 100
The composition and the weight part of VITAMIN are as follows:
Composition Mg/L
Vitamin H 0.0005
Choline chloride 60 20
D-calcium pantothenate 0.5
Folic acid 10
Inositol 3.0
Niacinamide 25
Pyridoxal hydrochloride 0.1
Pyridoxine hydrochloride 100
Riboflavin 0.1
The plain hydrochloride 3 of nuclear ammonia
Vitamin B12 0.001
Vitamins C 0.2
Vitamin K3 0.001
Nicotinic acid 0.3
Vitamin A acetic acid 0.009
Vitamin D2 0.2
The composition and the weight part of inorganic salt are as follows:
Composition Mg/L
Calcium chloride 45
Magnesium chloride 65
Sal epsom 20
Repone K 800
Sodium-chlor 2500
Sodium dihydrogen phosphate-water 150
Sodium phosphate dibasic 25
Sodium bicarbonate 5000
The composition and the weight part of other nutritive substances are as follows:
Composition Mg/L
Glucose 2000
Phenol red 15
Sodium.alpha.-ketopropionate 60
Xanthine 4.5
Linolic acid 0.001
Thioctic Acid 0.8
Putrescine acid 0.001
Gsh 1.5
Thymidine 0.08
4-hydroxyethyl piperazine ethanesulfonic acid 3000
Thanomin 0.4.
Component that trace element comprises and content are:
Component Mg/L
Cupric sulfate pentahydrate 0.02
Sodium Fluoride 0.0008
Nine water Sodium Selenites 0.2
Barium acetate 0.001
Eight water Cadmium Sulphates 0.004
Zirconium oxychloride 0.001.
Component and content that Transferrin Transferrins,iron complexes and Insulin Regular Insulin albumen surrogate comprise are:
Composition Mg/L
Zinc sulfate 15
1,2-arachidonic acyl Yelkin TTS 1.0
Ferrous sulfate seven water 0.12
Iron nitrate nine water water 0.01
Ironic oxalate 2.0
Ethylenediamine tetraacetic acid (EDTA) 4 sodium 0.01.
Embodiment 5
Substratum is made up of minimum medium, trace element and Transferrin Transferrins,iron complexes and Insulin insulin substitution thing, and wherein, the composition and the composition of minimum medium are as follows:
Amino acid whose composition and weight part are as follows:
Composition Mg/L
Glycine 30
L-L-Ala 3
L-arginine monohydrochloride 450
One water L-amino-succinamic acid 3.0
L-aspartic acid 25.0
L-cysteine hydrochloride 3.0
One water L-cysteine hydrochloride 65.0
L-L-glutamic acid 4
One water L-L-Histidine hydrochloride 75
L-Isoleucine 20
L-leucine 150
L lysine HCL 50
L-methionine(Met) 40
L-phenylalanine 1.0
L-proline(Pro) 85
L-Serine 3.0
L-Threonine 30
L-tryptophane 0.5
L-tyrosine disodium two water 25
L-Xie Ansuan 15
The composition and the weight part of VITAMIN are as follows:
Composition Mg/L
Vitamin H 0.09
Choline chloride 60 3.0
D-calcium pantothenate 9
Folic acid 0.5
Inositol 35
Niacinamide 3.0
Pyridoxal hydrochloride 5
Pyridoxine hydrochloride 10
Riboflavin 3
The plain hydrochloride 0.1 of nuclear ammonia
Vitamin B12 0.9
Vitamins C 0.001
Vitamin K3 0.2
Nicotinic acid 0.008
Vitamin A acetic acid 0.3
Vitamin D2 0.001
The composition and the weight part of inorganic salt are as follows:
Composition Mg/L
Calcium chloride 250
Magnesium chloride 10
Sal epsom 100
Repone K 100
Sodium-chlor 10000
Sodium dihydrogen phosphate-water 25
Sodium phosphate dibasic 150
Sodium bicarbonate 900
The composition and the weight part of other nutritive substances are as follows:
Composition Mg/L
Glucose 6500
Phenol red 0.5
Sodium.alpha.-ketopropionate 300
Xanthine 0.5
Linolic acid 0.5
Thioctic Acid 0.005
Putrescine acid 0.25
Gsh 0.1
Thymidine 0.85
4-hydroxyethyl piperazine ethanesulfonic acid 3000
Thanomin 0.4.
Component that trace element comprises and content are:
Component Mg/L
Cupric sulfate pentahydrate 0.02
Sodium Fluoride 0.0008
Nine water Sodium Selenites 0.2
Barium acetate 0.001
Eight water Cadmium Sulphates 0.004
Zirconium oxychloride 0.001.
Component and content that Transferrin Transferrins,iron complexes and Insulin Regular Insulin albumen surrogate comprise are:
Composition Mg/L
Zinc sulfate 15
1,2-arachidonic acyl Yelkin TTS 1.0
Ferrous sulfate seven water 0.12
Iron nitrate nine water water 0.01
Ironic oxalate 2.0
Ethylenediamine tetraacetic acid (EDTA) 4 sodium 0.01.
This Chinese hamster ovary cell serum free medium can directly be replaced and contains the conventional substratum of serum, and Chinese hamster ovary cell does not need adaptive process.

Claims (10)

1. a Chinese hamster ovary cell amplifying protein-free substratum is characterized in that: comprise minimum medium, also comprise: trace element and Transferr in Transferrins,iron complexes and Insulin insulin substitution thing.
2. substratum as claimed in claim 1 is characterized in that: wherein said minimum medium comprises: amino acid, VITAMIN, inorganic salt, other nutritive substances.
3. substratum as claimed in claim 2 is characterized in that: component that wherein said minimum medium comprises and content are:
Amino acid whose composition and weight part are as follows:
Composition Mg/L
Glycine 2-30
L-L-Ala 3-40
L-arginine monohydrochloride 50-450
One water L-amino-succinamic acid 3.0-45.0
L-aspartic acid 5.0-2 5.0
L-cysteine hydrochloride 3.0-45.0
One water L-cysteine hydrochloride 10.0-65.0
L-L-glutamic acid 4-35
One water L-L-Histidine hydrochloride 10-7 5
L-Isoleucine 20-100
L-leucine 15-150
L lysine HCL 50-300
L-methionine(Met) 3-40
L-phenylalanine 1.0-15
L-proline(Pro) 15-85
L-Serine 3.0-30
L-Threonine 3.0-30
L-tryptophane 0.5-8
L-tyrosine disodium two water 3.0-25
L-Xie Ansuan 15-100
The composition and the weight part of VITAMIN are as follows:
Composition Mg/L
Vitamin H 0.0005-0.09
Choline chloride 60 3.0-20
D-calcium pantothenate 0.5-9
Folic acid 0.5-10
Inositol 3.0-35
Niacinamide 3.0-25
Pyridoxal hydrochloride 0.1-5
Pyridoxine hydrochloride 10-100
Riboflavin 0.1-3
The plain hydrochloride 0.1-3 of nuclear ammonia
Vitamin B12 0.001-0.9
Vitamins C 0.001-0.2
Vitamin K3 0.001-0.2
Nicotinic acid 0.008-0.3
Vitamin A acetic acid 0.009-0.3
Vitamin D2 0.001-0.2
The composition and the weight part of inorganic salt are as follows:
Composition Mg/L
Calcium chloride 45-250
Magnesium chloride 10-65
Sal epsom 20-100
Repone K 100-800
Sodium-chlor 2500-10000
Sodium dihydrogen phosphate-water 25-150
Sodium phosphate dibasic 25-150
Sodium bicarbonate 900-5000
The composition and the weight part of other nutritive substances are as follows:
Composition Mg/L
Glucose 2000-6500
Phenol red 0.5-15
Sodium.alpha.-ketopropionate 60-300
Xanthine 0.5-4.5
Linolic acid 0.001-0.5
Thioctic Acid 0.005-0.8
Putrescine acid 0.001-0.25
Gsh 0.1-1.5
Thymidine 0.08-0.85
4-hydroxyethyl piperazine ethanesulfonic acid 800-3000
Thanomin 0.4-4.0.
4. substratum as claimed in claim 3 is characterized in that: component that wherein said minimum medium comprises and content are:
Amino acid whose composition and weight part are as follows:
Composition Mg/L
Glycine 7.5
L-L-Ala 8.9
L-arginine monohydrochloride 211
One water L-amino-succinamic acid 15.01
L-aspartic acid 13.3
L-cysteine hydrochloride 17.56
One water L-cysteine hydrochloride 31.29
L-L-glutamic acid 7.35
One water L-L-Histidine hydrochloride 31.48
L-Isoleucine 54.47
L-leucine 59.05
L lysine HCL 91.25
L-methionine(Met) 17.24
L-phenylalanine 5
L-proline 3 4.5
L-Serine 10.5
L-Threonine 11.9
L-tryptophane 2.04
L-tyrosine disodium two water 7.81
L-Xie Ansuan 46
The composition and the weight part of VITAMIN are as follows:
Composition Mg/L
Vitamin H 0.0035
Choline chloride 60 8.98
D-calcium pantothenate 2.24
Folic acid 2.65
Inositol 18
Niacinamide 12
Pyridoxal hydrochloride 1
Pyridoxine hydrochloride 35
Riboflavin 1
The plain hydrochloride 1 of nuclear ammonia
Vitamin B12 0.01
Vitamins C 0.05
Vitamin K3 0.05
Nicotinic acid 0.025
Vitamin A acetic acid 0.025
Vitamin D2 0.01
The composition and the weight part of inorganic salt are as follows:
Composition Mg/L
Calcium chloride 116.6
Magnesium chloride 28.64
Sal epsom 48.84
Repone K 311.8
Sodium-chlor 6995.5
Sodium dihydrogen phosphate-water 62.5
Sodium phosphate dibasic 71.02
Sodium bicarbonate 2200
The composition and the weight part of other nutritive substances are as follows:
Composition Mg/L
Glucose 3900
Phenol red 4
Sodium.alpha.-ketopropionate 150
Xanthine 1.66
Linolic acid 0.035
Thioctic Acid 0.075
Putrescine acid 0.004
Gsh 0.6
Thymidine 0.28
4-hydroxyethyl piperazine ethanesulfonic acid 1800
Thanomin 1.2.
5. substratum as claimed in claim 1 is characterized in that: component that described trace element comprises and content are:
Component Mg/L
Cupric sulfate pentahydrate 0.0009-0.02
Sodium Fluoride 0.0008-0.01
Nine water Sodium Selenite 0.05-0.2
Barium acetate 0.001-0.03
Eight water Cadmium Sulphate 0.0007-0.004
Zirconium oxychloride 0.001-0.0085.
6. substratum as claimed in claim 5 is characterized in that: component that described trace element comprises and content are:
Component Mg/L
Cupric sulfate pentahydrate 0.00182
Sodium Fluoride 0.00197
Nine water Sodium Selenites 0.094
Barium acetate 0.0079
Eight water Cadmium Sulphates 0.00122
Zirconium oxychloride 0.0015.
7. substratum as claimed in claim 1 is characterized in that: component and content that described Transferrin Transferrins,iron complexes and Insulin Regular Insulin albumen surrogate comprise are:
Composition Mg/L
Zinc sulfate 1.0-15
1,2-arachidonic acyl Yelkin TTS 1.0-16
Ferrous sulfate seven water 0.05-0.12
Iron nitrate nine water water 0.01-0.45
Ironic oxalate 0.1-2.0
Ethylenediamine tetraacetic acid (EDTA) 4 sodium 0.01-0.45.
8. substratum as claimed in claim 7 is characterized in that: component and content that described Transferrin Transferrins,iron complexes and Insulin Regular Insulin albumen surrogate comprise are:
Composition Mg/L
Zinc sulfate 5
1,2-arachidonic acyl Yelkin TTS 5
Ferrous sulfate seven water 0.094
Iron nitrate nine water water 0.05
Ironic oxalate 0.6
Ethylenediamine tetraacetic acid (EDTA) 4 sodium 0.03.
9. as the purposes of any described substratum among the claim 1-8 in cultivating Chinese hamster ovary cell.
10. purposes as claimed in claim 9 is characterized in that: described substratum, and this Chinese hamster ovary cell amplification serum free medium can directly be replaced and contains the conventional substratum of serum, and Chinese hamster ovary cell does not need adaptive process.
CNB2007103070769A 2007-12-28 2007-12-28 Gonad cell amplification culture medium of Chinese hamster and uses thereof Expired - Fee Related CN100506977C (en)

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CN101418330B (en) * 2008-06-20 2012-01-04 华东理工大学 Non protein culture medium adapted to large-scale culture of NSO cell and production of antibody
CN102653729A (en) * 2011-03-04 2012-09-05 上海赛金生物医药有限公司 Culture medium used for Chinese hamster ovary cells
CN102994441A (en) * 2012-09-19 2013-03-27 上海瀚康生物医药科技有限公司 Cell culture medium, and preparation method and use thereof
CN104073463A (en) * 2013-03-29 2014-10-01 上海中信国健药业股份有限公司 Serum-free protein-free culture medium supporting CHO (Chinese Hamster Ovary Cell) high density suspension culture
CN104212768A (en) * 2014-09-18 2014-12-17 中国科学院大连化学物理研究所 Protein-free culture medium for culturing microencapsulated recombinant Chinese hamster ovary (CHO) cells and preparation method thereof
CN105200006A (en) * 2015-10-30 2015-12-30 连云港市产品质量监督检验中心 Enhancer for improving culture medium for human hepatic cell toxicity test
CN109988741A (en) * 2019-04-10 2019-07-09 河南普诺易生物制品研究院有限公司 A kind of cell culture serum substitute and preparation method thereof, cell culture blood serum substituting composition, cell culture medium
CN115537378A (en) * 2022-11-24 2022-12-30 天信和(苏州)生物科技有限公司 Method for performing suspension culture on HEK293 cells and culture medium thereof
CN115627250A (en) * 2022-12-01 2023-01-20 天信和(苏州)生物科技有限公司 Serum-free medium for culturing insect cells and application thereof

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CN101418330B (en) * 2008-06-20 2012-01-04 华东理工大学 Non protein culture medium adapted to large-scale culture of NSO cell and production of antibody
CN102653729A (en) * 2011-03-04 2012-09-05 上海赛金生物医药有限公司 Culture medium used for Chinese hamster ovary cells
CN102653729B (en) * 2011-03-04 2014-07-16 上海赛金生物医药有限公司 Culture medium used for Chinese hamster ovary cells
CN102994441A (en) * 2012-09-19 2013-03-27 上海瀚康生物医药科技有限公司 Cell culture medium, and preparation method and use thereof
CN104073463A (en) * 2013-03-29 2014-10-01 上海中信国健药业股份有限公司 Serum-free protein-free culture medium supporting CHO (Chinese Hamster Ovary Cell) high density suspension culture
CN104073463B (en) * 2013-03-29 2019-01-01 三生国健药业（上海）股份有限公司 A kind of Serum-free and protein-free medium for supporting CHO high density suspension culture
CN104212768A (en) * 2014-09-18 2014-12-17 中国科学院大连化学物理研究所 Protein-free culture medium for culturing microencapsulated recombinant Chinese hamster ovary (CHO) cells and preparation method thereof
CN105200006A (en) * 2015-10-30 2015-12-30 连云港市产品质量监督检验中心 Enhancer for improving culture medium for human hepatic cell toxicity test
CN109988741A (en) * 2019-04-10 2019-07-09 河南普诺易生物制品研究院有限公司 A kind of cell culture serum substitute and preparation method thereof, cell culture blood serum substituting composition, cell culture medium
CN115537378A (en) * 2022-11-24 2022-12-30 天信和(苏州)生物科技有限公司 Method for performing suspension culture on HEK293 cells and culture medium thereof
CN115627250A (en) * 2022-12-01 2023-01-20 天信和(苏州)生物科技有限公司 Serum-free medium for culturing insect cells and application thereof
CN115627250B (en) * 2022-12-01 2023-04-04 天信和(苏州)生物科技有限公司 Serum-free medium for culturing insect cells and application thereof

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