CN110616183A - Low-serum culture medium for Vero cell culture and corresponding virus production - Google Patents

Low-serum culture medium for Vero cell culture and corresponding virus production Download PDF

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CN110616183A
CN110616183A CN201910900405.3A CN201910900405A CN110616183A CN 110616183 A CN110616183 A CN 110616183A CN 201910900405 A CN201910900405 A CN 201910900405A CN 110616183 A CN110616183 A CN 110616183A
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hydrochloride
sodium
monohydrate
acid
chloride
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齐智
孟丹丹
刘海英
房圆瑗
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Shandong Oracle Biotechnology Co Ltd
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Shandong Oracle Biotechnology Co Ltd
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    • C12N2770/24011—Flaviviridae
    • C12N2770/24311—Pestivirus, e.g. bovine viral diarrhea virus

Abstract

The invention discloses a low serum culture medium for Vero cell culture and corresponding virus production, which comprises the following components: amino acids, vitamins, inorganic salts, proteins and auxiliary components, wherein the amino acids comprise glycine, arginine hydrochloride, glutamine, lysine hydrochloride, serine, tyrosine disodium salt dihydrate and valine; the protein comprises full-component albumin, hydrolyzed milk protein (LAH), saturated iron human transferrin, full-chain recombinant insulin, and 4-hydroxyethyl piperazine ethanesulfonic acid (HEPES); the culture medium prepared by the invention is used for producing the vaccine of the Vero cell amplified porcine epidemic diarrhea virus, has simple formula components, low cost and easy acquisition, effectively improves the culture effect of the cells, and simultaneously keeps the good growth state of the cells.

Description

Low-serum culture medium for Vero cell culture and corresponding virus production
Technical Field
The invention relates to the field of production of biopharmaceutical vaccines, in particular to a low-serum culture medium for Vero cell culture and production of corresponding viruses.
Background
Vero cells are virus vaccine production cell lines approved by the world health organization and the Chinese biological product code. The virus vaccines produced by adopting the vero cell culture technology mainly comprise rabies vaccines, poliomyelitis vaccines, influenza vaccines, encephalitis B vaccines, hemorrhagic fever vaccines, hand-foot-and-mouth disease vaccines and the like. Vero cells have adherence characteristics, and the production of virus vaccines is largely determined to be realized by microcarrier immobilized culture of Vero cells.
However, the traditional Vero cell culture method is that 10% calf serum is added into an MEM culture medium to culture cells, the serum consumption is large, and the cost is high. The conditions of slow cell growth, cell death and the like can occur when the serum is reduced for culture. Part of commercially available low-serum culture media cannot meet the production requirements, and cannot keep the excellent growth state of cells under the low-serum culture condition, and meanwhile, the toxic value of the vaccine production semi-finished product is kept stable. The Vero cell amplified porcine epidemic diarrhea virus has two production modes, the pancreatin adding process is more critical to a cell culture medium, most of the commercially available low-serum culture media cannot meet the requirements, and based on the requirement, a culture medium for Vero cell low-serum culture is provided.
Disclosure of Invention
The invention aims to provide a low-serum culture medium for Vero cell culture and corresponding virus production, so as to solve the problems in the background technology.
In order to achieve the purpose, the invention provides the following technical scheme:
a low serum culture medium for Vero cell culture and corresponding virus production comprises the following components: amino acid, vitamin, inorganic salt, protein and auxiliary components, wherein the amino acid comprises 15-25mg/L of glycine, 5-10mg/L of alanine, 200-270mg/L of arginine hydrochloride, 50-60mg/L of asparagine monohydrate, 8-11mg/L of aspartic acid, 5-10mg/L of cysteine monohydrate, 50-70mg/L of cystine dihydrochloride, 8-15mg/L of glutamic acid, 400mg/L of glutamine, 25-50mg/L of histidine monohydrochloride, 60-75mg/L of isoleucine, 60-75mg/L of leucine, 80-110mg/L of lysine hydrochloride, 50-75mg/L of methionine, 40-50mg/L of phenylalanine, etc, 10-15mg/L of proline, 80-110mg/L of serine, 50-75mg/L of threonine, 10-15mg/L of tryptophan, 250mg/L of tyrosine disodium salt dihydrate, 50-75mg/L of valine, 8-12mg/L of hydroxyproline, 20-30mg/L of ornithine, 1-5mg/L of sodium ascorbyl phosphonate and 0.1-0.5mg/L of biotin; the protein comprises 500mg/L of total albumin 200-; the auxiliary component comprises D-glucose
4000mg/L of 2000-plus, 1-5mg/L of ethanolamine, 1-5mg/L of hypoxanthine sodium, 0.1-0.5mg/L of linoleic acid, 5-10mg/L of phenol red, 1-5mg/L of putrescine dihydrochloride, 100-120mg/L of sodium pyruvate, 100-300mg/L of galactose, 10-15mg/L of adenine, 100-200mg/L of fructose, 500-800mg/L of trehalose and 0.5-1mg/L of thymidine.
As a further scheme of the invention: the vitamins include choline chloride 5-7mg/L, D-calcium pantothenate 1-5mg/L, folic acid 1-5mg/L, nicotinamide 1-5mg/L, nicotinic acid 0.005-0.01mg/L, pyridoxine hydrochloride 1-5mg/L, pyridoxal hydrochloride 0.02-0.08mg/L, riboflavin 0.1-0.5mg/L, thiamine hydrochloride 10-15mg/L, vitamin B121-5mg/L, i-inositol 8-10mg/L, and vitamin A acetate 1-5 mg/L.
As a still further scheme of the invention: the inorganic salts comprise 150mg/L of anhydrous calcium chloride 120-.
As a still further scheme of the invention: 22.38mg/L of glycine, 6.48mg/L of alanine, 225.25mg/L of arginine hydrochloride, 59.00mg/L of asparagine monohydrate, 10.08mg/L of aspartic acid, 8.78mg/L of cysteine hydrochloride monohydrate, 69.65mg/L of cystine dihydrochloride, 10.43mg/L of glutamic acid, 532.50mg/L of glutamine, 34.74mg/L of histidine hydrochloride monohydrate, 71.24mg/L of isoleucine, 72.03mg/L of leucine, 105.63mg/L of lysine hydrochloride, 71.62mg/L of methionine, 46.74mg/L of phenylalanine, 13.88mg/L of proline, 106.63mg/L of serine, 67.23mg/L of threonine, 11.76mg/L of tryptophan, 310.40mg/L of disodium tyrosine dihydrate, 65.68mg/L of valine, 9.95mg/L of hydroxyproline, 25.00mg/L of ornithine, 2.50mg/L sodium ascorbyl phosphonate, 0.33mg/L biotin, 6.24mg/L, D-calcium pantothenate 4.87mg/L choline chloride, 3.08mg/L folic acid, 2.76mg/L nicotinamide, 0.01mg/L nicotinic acid, 1.11mg/L pyridoxine hydrochloride, 0.06mg/L pyridoxal hydrochloride, 0.23mg/L riboflavin, 10.25mg/L thiamine hydrochloride, 9.45mg/L vitamin B121.46mg/L, i-inositol, 1.03mg/L vitamin A acetate, 148.30mg/L anhydrous calcium chloride, 70.42mg/L anhydrous magnesium sulfate, 345.90mg/L potassium chloride, 2569.00mg/L sodium bicarbonate, 6797.75mg/L sodium chloride, 96.25mg/L sodium dihydrogen phosphate monohydrate, 35.51mg/L anhydrous disodium hydrogen phosphate, 1.11mg/L ferrous sulfate heptahydrate, 51.82mg/L of anhydrous magnesium chloride, 0.32mg/L, D of zinc sulfate heptahydrate-2175.50 mg/L of grape, 4.30mg/L of ethanolamine, 1.40mg/L of sodium hypoxanthine, 0.10mg/L of linoleic acid, 8.00mg/L of phenol red, 2.79mg/L of putrescine dihydrochloride, 110.00mg/L of sodium pyruvate, 200.00mg/L of galactose, 11.25mg/L of adenine, 100.00mg/L of fructose, 600.00mg/L of trehalose, 0.80mg/L of thymidine, 400mg/L of full-component albumin, 500mg/L of hydrolyzed milk protein (LAH), 10mg/L of iron human transferrin, 10mg/L of recombinant insulin, and 1400mg/L of 4-hydroxyethyl piperazine ethanesulfonic acid (HEPES).
As a still further scheme of the invention: the full component albumin is derived from bovine serum.
As a still further scheme of the invention: the human transferrin is saturated iron human transferrin.
As a still further scheme of the invention: the recombinant insulin is a full-chain recombinant insulin.
Compared with the prior art, the invention has the beneficial effects that:
the culture medium prepared by the invention is used for producing the vaccine of the Vero cell amplified porcine epidemic diarrhea virus, has simple formula components, low cost and easy acquisition, effectively improves the culture effect of the cells, and simultaneously keeps the good growth state of the cells.
Drawings
FIG. 1 shows the growth state of Vero cells in M199-HH medium in culture medium experiments.
FIG. 2 shows the M199-HH medium passage data of Vero cells in the culture medium experiment.
FIG. 3 shows the proliferation rate of Vero cells M199-HH medium in the medium experiment.
Detailed Description
The technical solution of the present invention will be described in further detail with reference to specific embodiments.
Example 1
A low serum culture medium for Vero cell culture and corresponding virus production comprises the following components: 22.38mg/L of glycine, 6.48mg/L of alanine, 225.25mg/L of arginine hydrochloride, 59.00mg/L of asparagine monohydrate, 10.08mg/L of aspartic acid, 8.78mg/L of cysteine hydrochloride monohydrate, 69.65mg/L of cystine dihydrochloride, 10.43mg/L of glutamic acid, 532.50mg/L of glutamine, 34.74mg/L of histidine hydrochloride monohydrate, 71.24mg/L of isoleucine, 72.03mg/L of leucine, 105.63mg/L of lysine hydrochloride, 71.62mg/L of methionine, 46.74mg/L of phenylalanine, 13.88mg/L of proline, 106.63mg/L of serine, 67.23mg/L of threonine, 11.76mg/L of tryptophan, 310.40mg/L of disodium tyrosine dihydrate, 65.68mg/L of valine, 9.95mg/L of hydroxyproline, 25.00mg/L of ornithine, 2.50mg/L sodium ascorbyl phosphonate, 0.33mg/L biotin, 6.24mg/L, D-calcium pantothenate 4.87mg/L choline chloride, 3.08mg/L folic acid, 2.76mg/L nicotinamide, 0.01mg/L nicotinic acid, 1.11mg/L pyridoxine hydrochloride, 0.06mg/L pyridoxal hydrochloride, 0.23mg/L riboflavin, 10.25mg/L thiamine hydrochloride, 9.45mg/L vitamin B121.46mg/L, i-inositol, 1.03mg/L vitamin A acetate, 148.30mg/L anhydrous calcium chloride, 70.42mg/L anhydrous magnesium sulfate, 345.90mg/L potassium chloride, 2569.00mg/L sodium bicarbonate, 6797.75mg/L sodium chloride, 96.25mg/L sodium dihydrogen phosphate monohydrate, 35.51mg/L anhydrous disodium hydrogen phosphate, 1.11mg/L ferrous sulfate heptahydrate, 51.82mg/L of anhydrous magnesium chloride, 0.32mg/L, D of zinc sulfate heptahydrate-2175.50 mg/L of grape, 4.30mg/L of ethanolamine, 1.40mg/L of sodium hypoxanthine, 0.10mg/L of linoleic acid, 8.00mg/L of phenol red, 2.79mg/L of putrescine dihydrochloride, 110.00mg/L of sodium pyruvate, 200.00mg/L of galactose, 11.25mg/L of adenine, 100.00mg/L of fructose, 600.00mg/L of trehalose, 0.80mg/L of thymidine, 400mg/L of full-component albumin, 500mg/L of hydrolyzed milk protein (LAH), 10mg/L of saturated iron human transferrin, 10mg/L of full-chain recombinant insulin, and 1400mg/L of 4-hydroxyethyl piperazine ethanesulfonic acid (HEPES).
Wherein the whole albumin is derived from bovine serum; the human transferrin is saturated iron human transferrin; the recombinant insulin is a full-chain recombinant insulin.
Example 2
A low serum culture medium for Vero cell culture and corresponding virus production comprises the following components: 15mg/L glycine, 5mg/L alanine, 200mg/L arginine hydrochloride, 50mg/L asparagine monohydrate, 8mg/L aspartic acid, 5mg/L cysteine hydrochloride monohydrate, 50mg/L cystine dihydrochloride, 8mg/L glutamic acid, 400mg/L glutamine, 25mg/L histidine hydrochloride monohydrate, 60mg/L isoleucine, 60mg/L leucine, 80mg/L lysine hydrochloride, 50mg/L methionine, 40mg/L phenylalanine, 10mg/L proline, 80mg/L serine, 50mg/L threonine, 10mg/L tryptophan, 250mg/L tyrosine disodium salt dihydrate, 50mg/L valine, 8mg/L hydroxyproline, 20mg/L ornithine, 200mg/L valine, or arginine hydrochloride, 1mg/L sodium ascorbyl phosphonate, 0.1mg/L biotin, 1mg/L choline chloride, 5mg/L, D-calcium pantothenate, 1mg/L folic acid, 1mg/L nicotinamide, 0.005mg/L nicotinic acid, 1mg/L pyridoxine hydrochloride, 0.02mg/L pyridoxal hydrochloride, 0.1mg/L riboflavin, 10mg/L thiamine hydrochloride, 8mg/L vitamin B121 mg/L, i-inositol, 1mg/L vitamin A acetate, 120mg/L anhydrous calcium chloride, 50mg/L anhydrous magnesium sulfate, 200mg/L potassium chloride, 2000mg/L sodium bicarbonate, 5000mg/L sodium chloride, 80mg/L sodium dihydrogen phosphate monohydrate, 30mg/L anhydrous disodium hydrogen phosphate, 1mg/L ferrous sulfate heptahydrate, 1mg/L sodium hydrogen phosphate, 40mg/L of anhydrous magnesium chloride, 0.1mg/L, D of zinc sulfate heptahydrate, 2000mg/L of glucose, 1mg/L of ethanolamine, 1mg/L of hypoxanthine sodium, 0.1mg/L of linoleic acid, 5mg/L of phenol red, 1mg/L of putrescine dihydrochloride, 100mg/L of sodium pyruvate, 100mg/L of galactose, 10mg/L of adenine, 100mg/L of fructose, 500mg/L of trehalose, 0.5mg/L of thymidine, 200mg/L of full-component albumin, 300mg/L of lactoprotein hydrolysate (LAH), 5mg/L of saturated ferric transferrin, 5mg/L of full-chain recombinant insulin and 1200mg/L of 4-hydroxyethyl piperazine ethanesulfonic acid (HEPES).
Wherein the whole albumin is derived from bovine serum; the human transferrin is saturated iron human transferrin; the recombinant insulin is a full-chain recombinant insulin.
Example 3
A low serum culture medium for Vero cell culture and corresponding virus production comprises the following components: 25mg/L glycine, 10mg/L alanine, 270mg/L arginine hydrochloride, 60mg/L asparagine monohydrate, 11mg/L aspartic acid, 10mg/L cysteine hydrochloride monohydrate, 70mg/L cystine dihydrochloride, 15mg/L glutamic acid, 600mg/L glutamine, 50mg/L histidine hydrochloride monohydrate, 75mg/L isoleucine, 75mg/L leucine, 110mg/L lysine hydrochloride, 75mg/L methionine, 50mg/L phenylalanine, 15mg/L proline, 110mg/L serine, 75mg/L threonine, 15mg/L tryptophan, 320mg/L tyrosine disodium salt dihydrate, 75mg/L valine, 12mg/L hydroxyproline, 30mg/L ornithine, 5mg/L sodium ascorbyl phosphonate, 0.5mg/L biotin, 7mg/L, D-calcium pantothenate, 5mg/L choline chloride, 5mg/L folic acid, 5mg/L nicotinamide, 0.01mg/L nicotinic acid, 5mg/L pyridoxine hydrochloride, 0.08mg/L pyridoxal hydrochloride, 0.5mg/L riboflavin, 15mg/L thiamine hydrochloride, 10mg/L vitamin B125 mg/L, i-inositol, 5mg/L vitamin A acetate, 150mg/L anhydrous calcium chloride, 80mg/L anhydrous magnesium sulfate, 400mg/L potassium chloride, 3000mg/L sodium bicarbonate, 7000mg/L sodium chloride, 100mg/L sodium dihydrogen phosphate monohydrate, 40mg/L anhydrous disodium hydrogen phosphate, 5mg/L ferrous sulfate heptahydrate, 5mg/L sodium hydrogen phosphate, 55mg/L of anhydrous magnesium chloride, 0.5mg/L, D of zinc sulfate heptahydrate, 4000mg/L of glucose, 5mg/L of ethanolamine, 5mg/L of hypoxanthine sodium, 0.5mg/L of linoleic acid, 10mg/L of phenol red, 5mg/L of putrescine dihydrochloride, 120mg/L of sodium pyruvate, 300mg/L of galactose, 15mg/L of adenine, 200mg/L of fructose, 800mg/L of trehalose, 1mg/L of thymidine, 500mg/L of full-component albumin, 600mg/L of lactoprotein hydrolysate (LAH), 10mg/L of saturated iron human transferrin, 10mg/L of full-chain recombinant insulin and 1500mg/L of 4-hydroxyethyl piperazine ethanesulfonic acid (HEPES).
Wherein the whole albumin is derived from bovine serum; the human transferrin is saturated iron human transferrin; the recombinant insulin is a full-chain recombinant insulin.
Example 4
A low serum culture medium for Vero cell culture and corresponding virus production comprises the following components: 15mg/L glycine, 5mg/L alanine, 200mg/L arginine hydrochloride, 50mg/L asparagine monohydrate, 8mg/L aspartic acid, 5mg/L cysteine hydrochloride monohydrate, 50mg/L cystine dihydrochloride, 8mg/L glutamic acid, 400mg/L glutamine, 25mg/L histidine hydrochloride monohydrate, 60mg/L isoleucine, 60mg/L leucine, 80mg/L lysine hydrochloride, 50mg/L methionine, 40mg/L phenylalanine, 10mg/L proline, 80mg/L serine, 50mg/L threonine, 10mg/L tryptophan, 250mg/L tyrosine disodium salt dihydrate, 50mg/L valine, 8mg/L hydroxyproline, 20mg/L ornithine, 200mg/L valine, or arginine hydrochloride, Sodium ascorbyl phosphonate 1mg/L, biotin 0.1mg/L, choline chloride 6.24mg/L, D-calcium pantothenate 4.87mg/L, folic acid 3.08mg/L, nicotinamide 2.76mg/L, nicotinic acid 0.01mg/L, pyridoxine hydrochloride 1.11mg/L, pyridoxal hydrochloride 0.06mg/L, riboflavin 0.23mg/L, thiamine hydrochloride 10.25mg/L, vitamin B121.46mg/L, i-inositol 9.45mg/L, vitamin A acetate 1.03mg/L, anhydrous calcium chloride 148.30mg/L, anhydrous magnesium sulfate 70.42mg/L, potassium chloride 345.90mg/L, sodium bicarbonate 2569.00mg/L, sodium chloride 6797.75mg/L, monobasic sodium phosphate 96.25mg/L, anhydrous disodium hydrogen phosphate 35.51mg/L, ferrous sulfate heptahydrate 1.11mg/L, 51.82mg/L of anhydrous magnesium chloride, 0.32mg/L, D mg/L of zinc sulfate heptahydrate, 2000mg/L of glucose, 1mg/L of ethanolamine, 1mg/L of hypoxanthine sodium, 0.1mg/L of linoleic acid, 5mg/L of phenol red, 1mg/L of putrescine dihydrochloride, 100mg/L of sodium pyruvate, 100mg/L of galactose, 10mg/L of adenine, 100mg/L of fructose, 500mg/L of trehalose, 0.5mg/L of thymidine, 200mg/L of full-component albumin, 300mg/L of lactoprotein hydrolysate (LAH), 5mg/L of saturated ferritransferrin, 5mg/L of full-chain recombinant insulin and 1200mg/L of 4-hydroxyethyl piperazine ethanesulfonic acid (HEPES).
Wherein the whole albumin is derived from bovine serum; the human transferrin is saturated iron human transferrin; the recombinant insulin is a full-chain recombinant insulin.
Example 5
A low serum culture medium for Vero cell culture and corresponding virus production comprises the following components: 22.38mg/L of glycine, 6.48mg/L of alanine, 225.25mg/L of arginine hydrochloride, 59.00mg/L of asparagine monohydrate, 10.08mg/L of aspartic acid, 8.78mg/L of cysteine hydrochloride monohydrate, 69.65mg/L of cystine dihydrochloride, 10.43mg/L of glutamic acid, 532.50mg/L of glutamine, 34.74mg/L of histidine hydrochloride monohydrate, 71.24mg/L of isoleucine, 72.03mg/L of leucine, 105.63mg/L of lysine hydrochloride, 71.62mg/L of methionine, 46.74mg/L of phenylalanine, 13.88mg/L of proline, 106.63mg/L of serine, 67.23mg/L of threonine, 11.76mg/L of tryptophan, 310.40mg/L of disodium tyrosine dihydrate, 65.68mg/L of valine, 9.95mg/L of hydroxyproline, 25.00mg/L of ornithine, 2.50mg/L ascorbyl phosphonate sodium, 0.33mg/L biotin, 5mg/L, D-calcium pantothenate 1mg/L choline chloride, 1mg/L folic acid, 1mg/L nicotinamide, 0.005mg/L nicotinic acid, 1mg/L pyridoxine hydrochloride, 0.02mg/L pyridoxal hydrochloride, 0.1mg/L riboflavin, 10mg/L thiamine hydrochloride, vitamin B121 mg/L, i-inositol 8mg/L vitamin A acetate, 120mg/L anhydrous calcium chloride, 50mg/L anhydrous magnesium sulfate, 200mg/L potassium chloride, 2000mg/L sodium bicarbonate, 5000mg/L sodium chloride, 80mg/L sodium dihydrogen phosphate monohydrate, 30mg/L anhydrous disodium hydrogen phosphate, 1mg/L ferrous sulfate heptahydrate, 1mg/L sodium sulfate, 40mg/L of anhydrous magnesium chloride, 0.1mg/L, D of zinc sulfate heptahydrate, 4000mg/L of glucose, 5mg/L of ethanolamine, 5mg/L of hypoxanthine sodium, 0.5mg/L of linoleic acid, 10mg/L of phenol red, 5mg/L of putrescine dihydrochloride, 120mg/L of sodium pyruvate, 300mg/L of galactose, 15mg/L of adenine, 200mg/L of fructose, 800mg/L of trehalose, 1mg/L of thymidine, 500mg/L of full-component albumin, 600mg/L of lactoprotein hydrolysate (LAH), 10mg/L of saturated iron human transferrin, 10mg/L of full-chain recombinant insulin and 1500mg/L of 4-hydroxyethyl piperazine ethanesulfonic acid (HEPES).
Wherein the whole albumin is derived from bovine serum; the human transferrin is saturated iron human transferrin; the recombinant insulin is a full-chain recombinant insulin.
Experiment in culture Medium
The culture medium prepared by the invention is used for producing the vaccine of the Vero cell amplified porcine epidemic diarrhea virus, has simple formula components, low cost and easy acquisition, effectively improves the culture effect of the cells, and simultaneously keeps the good growth state of the cells.
While the preferred embodiments of the present invention have been described in detail, the present invention is not limited to the above embodiments, and various changes can be made without departing from the spirit of the present invention within the knowledge of those skilled in the art.

Claims (7)

1. A low serum culture medium for Vero cell culture and corresponding virus production is characterized by comprising the following components: amino acid, vitamin, inorganic salt, protein and auxiliary components, wherein the amino acid comprises 15-25mg/L of glycine, 5-10mg/L of alanine, 200-270mg/L of arginine hydrochloride, 50-60mg/L of asparagine monohydrate, 8-11mg/L of aspartic acid, 5-10mg/L of cysteine monohydrate, 50-70mg/L of cystine dihydrochloride, 8-15mg/L of glutamic acid, 400mg/L of glutamine, 25-50mg/L of histidine monohydrate, 60-75mg/L of isoleucine, 60-75mg/L of leucine, 80-110mg/L of lysine hydrochloride, 50-75mg/L of methionine, and the like, 40-50mg/L phenylalanine, 10-15mg/L proline, 80-110mg/L serine, 50-75mg/L threonine, 10-15mg/L tryptophan, 250-320mg/L tyrosine disodium salt dihydrate, 50-75mg/L valine, 8-12mg/L hydroxyproline, 20-30mg/L ornithine, 1-5mg/L sodium ascorbyl phosphonate and 0.1-0.5mg/L biotin; the protein comprises 500mg/L of whole albumin 200-; the auxiliary components comprise 4000mg/L of D-glucose 2000-.
2. The low serum medium for Vero cell culture and production of corresponding viruses according to claim 1, wherein said vitamins comprise choline chloride 5-7mg/L, D-calcium pantothenate 1-5mg/L, folic acid 1-5mg/L, nicotinamide 1-5mg/L, nicotinic acid 0.005-0.01mg/L, pyridoxine hydrochloride 1-5mg/L, pyridoxal hydrochloride 0.02-0.08mg/L, riboflavin 0.1-0.5mg/L, thiamine hydrochloride 10-15mg/L, vitamin B121-5mg/L, i-inositol 8-10mg/L, and vitamin A acetate 1-5 mg/L.
3. The low serum medium for Vero cell culture and corresponding virus production according to claim 2 wherein the inorganic salts comprise anhydrous calcium chloride 120-150mg/L, anhydrous magnesium sulfate 50-80mg/L, potassium chloride 200-400mg/L, sodium bicarbonate 2000-3000mg/L, sodium chloride 5000-7000mg/L, sodium dihydrogen phosphate monohydrate 80-100mg/L, anhydrous disodium hydrogen phosphate 30-40mg/L, ferrous sulfate heptahydrate 1-5mg/L, anhydrous magnesium chloride 40-55mg/L, zinc sulfate heptahydrate 0.1-0.5 mg/L.
4. A low serum medium for Vero cell culture and corresponding virus production according to claim 3, comprising the following composition: 22.38mg/L of glycine, 6.48mg/L of alanine, 225.25mg/L of arginine hydrochloride, 59.00mg/L of asparagine monohydrate, 10.08mg/L of aspartic acid, 8.78mg/L of cysteine hydrochloride monohydrate, 69.65mg/L of cystine dihydrochloride, 10.43mg/L of glutamic acid, 532.50mg/L of glutamine, 34.74mg/L of histidine hydrochloride monohydrate, 71.24mg/L of isoleucine, 72.03mg/L of leucine, 105.63mg/L of lysine hydrochloride, 71.62mg/L of methionine, 46.74mg/L of phenylalanine, 13.88mg/L of proline, 106.63mg/L of serine, 67.23mg/L of threonine, 11.76mg/L of tryptophan, 310.40mg/L of disodium tyrosine dihydrate, 65.68mg/L of valine, or mixtures thereof, Hydroxyproline 9.95mg/L, ornithine 25.00mg/L, sodium ascorbyl phosphonate 2.50mg/L, biotin 0.33mg/L, choline chloride 6.24mg/L, D-calcium pantothenate 4.87mg/L, folic acid 3.08mg/L, nicotinamide 2.76mg/L, nicotinic acid 0.01mg/L, pyridoxine hydrochloride 1.11mg/L, pyridoxal hydrochloride 0.06mg/L, riboflavin 0.23mg/L, thiamine hydrochloride 10.25mg/L, vitamin B121.46mg/L, i-inositol 9.45mg/L, vitamin A acetate 1.03mg/L, anhydrous calcium chloride 148.30mg/L, anhydrous magnesium sulfate 70.42mg/L, potassium chloride 345.90mg/L, sodium bicarbonate 2569.00mg/L, sodium chloride 6797.75mg/L, 6797.75mg/L, 96.25mg/L of sodium dihydrogen phosphate monohydrate, 35.51mg/L of anhydrous disodium hydrogen phosphate, 1.11mg/L of ferrous sulfate heptahydrate, 51.82mg/L of anhydrous magnesium chloride, 0.32mg/L, D mg/L of zinc sulfate heptahydrate, 2175.50mg/L of grape, 4.30mg/L of ethanolamine, 1.40mg/L of hypoxanthine sodium, 0.10mg/L of linoleic acid, 8.00mg/L of phenol red, 2.79mg/L of putrescine dihydrochloride, 110.00mg/L of sodium pyruvate, 200.00mg/L of galactose, 11.25mg/L of adenine, 100.00mg/L of fructose, 600.00mg/L of trehalose, 0.80mg/L of thymidine, 400mg/L of full-component albumin, 500mg/L of hydrolyzed lactoprotein, 10mg/L of iron human transferrin, 10mg/L of recombinant insulin, 10mg/L of insulin, and the like, 1400mg/L of 4-hydroxyethyl piperazine ethanesulfonic acid.
5. A medium according to claim 4 for the low serum culture of Vero cells wherein the full component albumin is derived from bovine serum.
6. The medium for low serum culture of Vero cells according to claim 4 wherein the human transferrin is saturated iron human transferrin.
7. The medium for low serum culture of Vero cells according to claim 4 wherein the recombinant insulin is a full chain recombinant insulin.
CN201910900405.3A 2019-09-23 2019-09-23 Low-serum culture medium for Vero cell culture and corresponding virus production Pending CN110616183A (en)

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