US5326699A
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Serum-free medium for culturing animal cells
Assignee: Kurashiki Spinning Co Ltd
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Filing: 1993-09-14
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Granted: 1994-07-05
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Expires: 2013-09-14
Abstract
Abstract The present invention provides a serum-free medium for culturing animal cells characterized in that it comprises 8.0-14.0 mg/l of methionine. Although a wide variety of animal cells can effectively be cultured by means of the serum-free medium according to the present invention, said serum-free medium produces an excellent growth effect for culturing animal epithelial cells in particular.
Inventors
- 👤 Hisashi Torishima
- 👤 Hirokuni Arakawa
- 👤 Ryohei Yamamoto
- 👤 Toyokazu Nishino
- 👤 Chikaaki Sakai
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-11-04 12:25
CPC Classifications
C; C12; C12N; C12N5/00; C12N5/06; C12N5/0602; C12N5/0625; C12N5/0629; C12N5/0018; C12N5/0037; C12N5/0618; C12N5/0621; C12N2500/00; C12N2500/30; C12N2500/32
Claims Analysis
Total Claims: 22
Claim 1
Independent Claim
84 words
A serum-free medium for culturing animal epithelial cells comprising 9.0-14.0 mg/l of methionine, 5.0-11.5 mg/l of threonine, 7.0-11.0 mg/l of tyrosine, 9.0-14.5 mg/l of phenylalanine, 400.0-550.0 mg/l of glutamine, 7.0-9.5 mg/l of aspartic acid, 19.0-28.0 mg/l of lysine HCl, 33.0-40.0 mg/l of serine, and up to 0.1 mM of Ca in the form of calcium chloride; and one or more conventional ingredients selected from the group consisting of glucose, vitamins, minerals, growth factors, HEPES, and amino acids other than the foregoing listed amino acids.
Claim 2
Dependent Claim
84 words
A serum-free medium for culturing animal epithelial cells comprising 9.0-14.0 mg/l of methionine, 5.0-11.5 mg/l of threonine, 7.0-11.0 mg/l of tyrosine, 9.0-14.5 mg/l of phenylalanine, 400.0-550.0 mg/l of glutamine, 7.0-9.5 mg/l of aspartic acid, 19.0-28.0 mg/l of lysine HCl, 33.0-40.0 mg/l of serine, and up to 0.1 mM of Ca in the form of calcium chloride; and one or more conventional ingredients selected from the group consisting of glucose, vitamins, minerals, growth factors, HEPES, and amino acids other than the foregoing listed amino acids.
Claim 3
Dependent Claim
38 words
The serum-free medium of claim 1, wherein the other amino acids are selected from the group consisting of alanine, arginine, asparagine, cysteine, glutamic acid, glycine, histidine, isoleucine, leucine, proline, tryptophan, valine, and alkaline salts or acid salts thereof.
Claim 4
Dependent Claim
47 words
The serum-free medium of claim 1, wherein the vitamins are selected from the group consisting of biotin, folic acid, lipoic acid, nicotinamide, pantothenic acid, pyridoxine, riboflavin, thiamine, vitamin B 12 , adenine, choline chloride, inositol, hypoxanthine, putrescine, pyruvic acid, thymidine, and alkaline salts or acid salts thereof.
Claim 5
Dependent Claim
31 words
The serum-free medium of claim 1, wherein the minerals are selected from the group consisting of Ca, K, Mg, Na, Cu, Fe, Se, Mn, Si, Mo, V, Ni, Sn, and Zn.
Claim 6
Dependent Claim
26 words
The serum-free medium of claim 1, wherein the growth factors are selected from the group consisting of insulin, hydrocortisone, epidermal growth factor, and bovine pituitary extract.
Claim 7
Dependent Claim
14 words
The serum-free medium of claim 1, wherein the medium additionally contains phenol red (Na).
Claim 8
Dependent Claim
197 words
A serum free medium for culturing animal epithelial cells which comprises 9.0-14.0 mg/l of methionine, 5.0-11.5 mg/l of threonine, 7.0-11.0 mg/l of tyrosine, 9.0-14.5 mg/l of phenylalanine, 400.0-550.0 mg/l of glutamine, 7.0-9.5 mg/l of aspartic acid, 19.0-28.0 mg/l of lysine HCl, 33.0-40.0 mg/l of serine, 0.1 mM or less of calcium chloride; alanine, arginine HCl, asparagine H 2 O, cysteine HCl H 20 , glutamic acid, glycine, histidine HCl H 2 O, isoleucine, leucine, proline, tryptophan, valine, biotin, folic acid, lipoic acid, nicotinamide, calcium pantothenate, pyridoxine HCl, riboflavin, thiamine HCl, vitamin B 12 , adenine, choline chloride, inositol, hypoxanthine, putrescine HCl, sodium pyruvate, thymidine, KCl, MgCl 2 6H 2 O, NaCl, NaH 2 PO 4 7H 2 O, KH 2 PO 4 , CuSO 4 5H 2 O, FeSO 4 7H 2 O, Na 2 SeO 4 , MnSO 4 5H 2 O, Na 2 SiO 3 9H 2 O, (NH 4 ) 6 Mo 7 O 24 4H 2 O, NH 4 VO 3 , NiCl 2 6H 2 O, SnCl 2 2H 2 O, ZnSO 4 7H 2 O, NaHCO 3 glucose, HEPES, phenol red (Na), insulin, hydrocortisone, epidermal growth factor, and bovine pituitary extract.
Claim 9
Dependent Claim
197 words
A serum free medium for culturing animal epithelial cells which comprises 9.0-14.0 mg/l of methionine, 5.0-11.5 mg/l of threonine, 7.0-11.0 mg/l of tyrosine, 9.0-14.5 mg/l of phenylalanine, 400.0-550.0 mg/l of glutamine, 7.0-9.5 mg/l of aspartic acid, 19.0-28.0 mg/l of lysine HCl, 33.0-40.0 mg/l of serine, 0.1 mM or less of calcium chloride; alanine, arginine HCl, asparagine H 2 O, cysteine HCl H 20 , glutamic acid, glycine, histidine HCl H 2 O, isoleucine, leucine, proline, tryptophan, valine, biotin, folic acid, lipoic acid, nicotinamide, calcium pantothenate, pyridoxine HCl, riboflavin, thiamine HCl, vitamin B 12 , adenine, choline chloride, inositol, hypoxanthine, putrescine HCl, sodium pyruvate, thymidine, KCl, MgCl 2 6H 2 O, NaCl, NaH 2 PO 4 7H 2 O, KH 2 PO 4 , CuSO 4 5H 2 O, FeSO 4 7H 2 O, Na 2 SeO 4 , MnSO 4 5H 2 O, Na 2 SiO 3 9H 2 O, (NH 4 ) 6 Mo 7 O 24 4H 2 O, NH 4 VO 3 , NiCl 2 6H 2 O, SnCl 2 2H 2 O, ZnSO 4 7H 2 O, NaHCO 3 glucose, HEPES, phenol red (Na), insulin, hydrocortisone, epidermal growth factor, and bovine pituitary extract.
Claim 10
Dependent Claim
17 words
The serum-free medium of claim 1, wherein the epithelial cells are epidermal keratinocytes or corneal epithelial cells.
Claim 11
Dependent Claim
13 words
The serum-free medium of claim 1, in which the culture is primary culture.
Claim 12
Dependent Claim
14 words
The serum-free medium of claim 1, wherein the epithelial cells are corneal epithelial cells.
Claim 13
Independent Claim
92 words
A method of culturing animal corneal epithelial cells comprising culturing said cells in a serum-free medium comprising 9.0-14.0 mg/l of methionine, 5.0-11.5 mg/l of threonine, 7.0-11.0 mg/l of tyrosine, 9.0-14.5 mg/l of phenylalanine, 400.0-550.0 mg/l of glutamine, 7.0-9.5 mg/l of aspartic acid, 19.0-28.0 mg/l of lysine HCl, 33.0-40.0 mg/l of serine, and up to 0.1 mM of Ca in the form of calcium chloride; and one or more conventional ingredients selected from the group consisting of glucose, vitamins, minerals, growth factors, HEPES, and amino acids other than the foregoing listed amino acids.
Claim 14
Independent Claim
92 words
A method of culturing animal corneal epithelial cells comprising culturing said cells in a serum-free medium comprising 9.0-14.0 mg/l of methionine, 5.0-11.5 mg/l of threonine, 7.0-11.0 mg/l of tyrosine, 9.0-14.5 mg/l of phenylalanine, 400.0-550.0 mg/l of glutamine, 7.0-9.5 mg/l of aspartic acid, 19.0-28.0 mg/l of lysine HCl, 33.0-40.0 mg/l of serine, and up to 0.1 mM of Ca in the form of calcium chloride; and one or more conventional ingredients selected from the group consisting of glucose, vitamins, minerals, growth factors, HEPES, and amino acids other than the foregoing listed amino acids.
Claim 15
Dependent Claim
37 words
The method of claim 11, wherein the other amino acids are selected from the group consisting of alanine, arginine, asparagine, cysteine, glutamic acid, glycine, histidine, isoleucine, leucine, proline, tryptophan, valine, and alkaline salts or acid salts thereof.
Claim 16
Dependent Claim
46 words
The method of claim 11, wherein the vitamins are selected from the group consisting of biotin, folic acid, lipoic acid, nicotinamide, pantothenic acid, pyridoxine, riboflavin, thiamine, vitamin B 12 , adenine, choline chloride, inositol, hypoxanthine, putrescine, pyruvic acid, thymidine, and alkaline salts or acid salts thereof.
Claim 17
Dependent Claim
30 words
The method of claim 11, wherein the minerals are selected from the group consisting of Ca, K, Mg, Na, Cu, Fe, Se, Mn, Si, Mo, V, Ni, Sn, and Zn.
Claim 18
Dependent Claim
25 words
The method of claim 11, wherein the growth factors are selected from the group consisting of insulin, hydrocortisone, epidermal growth factor, and bovine pituitary extract.
Claim 19
Dependent Claim
13 words
The method of claim 11, wherein the medium additionally contains phenol red (Na).
Claim 20
Independent Claim
205 words
A method of culturing animal corneal epithelial cells which comprises culturing said cells in a serium-free medium comprising 9.0-14.0 mg/l of methionine, 5.0-11.5 mg/l of threonine, 7.0-11.0 mg/l of tyrosine, 9.0-14.5 mg/l of phenylalanine, 400.0-550.0 mg/l of glutamine, 7.0-9.5 mg/l of aspartic acid, 19.0-28.0 mg/l of lysine HCl, 33.0-40.0 mg/l of serine, 0.1 mM or less of calcium chloride; alanine, arginine HCl asparagine H 2 O, cysteine HCl H 2 O, glutamic acid, glycine, histidine HCl H 2 O, isoleucine, leucine, proline, tryptophan, valine, biotin, folic acid, lipoic acid, nicotinamide, calcium pantothenate, pyridoxine HCl, riboflavin, thiamine HCl, vitamin B 12 , adenine, choline chloride, inositol, hypoxanthine, putrescine HCl, sodium pyruvate, thymidine, KCl, MgCl 2 6H 2 O, NaCl, NaH 2 PO 4 7H 2 O, KH 2 PO 4 , CuSO 4 5H 2 O, FeSO 4 7H 2 O, Na 2 SeO 4 , MnSO 4 5H 2 O, Na 2 SiO 3 9H 2 O, (NH 4 ) 6 Mo 7 O 24 4H 2 O, NH 4 VO 3 , NiCl 2 6H 2 O, SnCl 2 2H 2 O, ZnSO 4 7H 2 O, NaHCO 3 , glucose, HEPES, phenol red (Na), insulin, hydrocortisone, epidermal growth factor, and bovine pituitary extract.
Claim 21
Independent Claim
205 words
A method of culturing animal corneal epithelial cells which comprises culturing said cells in a serium-free medium comprising 9.0-14.0 mg/l of methionine, 5.0-11.5 mg/l of threonine, 7.0-11.0 mg/l of tyrosine, 9.0-14.5 mg/l of phenylalanine, 400.0-550.0 mg/l of glutamine, 7.0-9.5 mg/l of aspartic acid, 19.0-28.0 mg/l of lysine HCl, 33.0-40.0 mg/l of serine, 0.1 mM or less of calcium chloride; alanine, arginine HCl asparagine H 2 O, cysteine HCl H 2 O, glutamic acid, glycine, histidine HCl H 2 O, isoleucine, leucine, proline, tryptophan, valine, biotin, folic acid, lipoic acid, nicotinamide, calcium pantothenate, pyridoxine HCl, riboflavin, thiamine HCl, vitamin B 12 , adenine, choline chloride, inositol, hypoxanthine, putrescine HCl, sodium pyruvate, thymidine, KCl, MgCl 2 6H 2 O, NaCl, NaH 2 PO 4 7H 2 O, KH 2 PO 4 , CuSO 4 5H 2 O, FeSO 4 7H 2 O, Na 2 SeO 4 , MnSO 4 5H 2 O, Na 2 SiO 3 9H 2 O, (NH 4 ) 6 Mo 7 O 24 4H 2 O, NH 4 VO 3 , NiCl 2 6H 2 O, SnCl 2 2H 2 O, ZnSO 4 7H 2 O, NaHCO 3 , glucose, HEPES, phenol red (Na), insulin, hydrocortisone, epidermal growth factor, and bovine pituitary extract.
Claim 22
Dependent Claim
17 words
The method of claim 17 further comprising the steps of subculturing the cells in the serum-free medium.
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