US4657866A
Expired / Lapsed
US
⏱️ 0.0 years remaining
Serum-free, synthetic, completely chemically defined tissue culture media
Assignee: Individual
•
Filing: 1984-09-13
•
Granted: 1987-04-14
•
Expires: 2004-09-13
Abstract
Abstract A serum-free, synthetic tissue culture media is described which is completely defined chemically. The two media described can be used for growing all types of human or animal cell lines in tissue culture without addition of any protein, amino acids, hormones, sources of energy, salts, vitamins, etc. with normally used procedures and methods. The media do not require any supplementation with fetal calf serum to support growth of neural cells.
Inventors
- 👤 Sudhir Kumar
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-11-05 21:21
CPC Classifications
C; C12; C12N; C12N5/00; C12N5/06; C12N5/0602; C12N5/0618; C12N5/0018; C12N5/005; C12N2500/00; C12N2500/30; C12N2500/32; C12N2500/38; C12N2500/40; C12N2500/90; C12N2500/95; C12N2501/00; C12N2501/30; C12N2501/38; C12N2501/39; Y; Y10; Y10S; Y10S435/00; Y10S435/8215; Y10S435/948
Claims Analysis
Total Claims: 13
Claim 1
Independent Claim
273 words
(a) A cell culturing medium comprising an amino acid composition containing: ______________________________________
Item mg/liter
______________________________________
Alanine 60
Alpha-Amino-n-butyric Acid
5
Arginine 150
Asparagine 5
Aspartic Acid 75
Cysteine-HCl 100
Cystine 25
Ergothionine 10
Glutamic Acid 500
Glutamine 800
Glycine 500
Histidine, HCl 50
Homocysteine 20
Isoleucine 75
Leucine 300
Lysine HCl 300
Methionine 50
Ornithine HCl 10
Phenylalanine 100
Proline 100
Serine 75
Taurine 50
Threonine 100
Tryptophane 50
Tyrosine 100
and Valine 200,
______________________________________ (b) a vitamin mixture containing: ______________________________________
Item mg/liter
______________________________________
DL-Alpha-Tocopherol
2.5
Ascorbic Acid 25
Biotin 20
Calciferol 2.5
Calcium-pentothenate
25
Choline Chloride 25
Cocarboxylase 20
Folic Acid 20
i-Inositol 15
Menadione 2.5
Niacin 5.0
Niacinamide 20
p-Aminobenzoic Acid
1.0
Pyridoxal HCl 20
Pyridoxine HCl 5.0
Riboflavin 20
Thiamine HCl 20
Vitamin B12 50
Vitamin A 10
and Citric Acid 20,
______________________________________ (c) other components containing: ______________________________________
Item mg/liter
______________________________________
Na--Acetate, 3 H.sub.2 O
100
ATP (Sodium Salt) 20
Estradiol 20
Cholate 25
Deoxyadenosine 15
Deoxyguanosine 15
Dextrose 1500
Flavin Adenine Dinucleotide
20
Galactosamine 120
Glucose-1-phosphate 25
Glucuronolactone 25
Hypoxanthine 100
5-Methyl-deoxycytidylic Acid
5
Phenol Red 15
Thioactic Acid 2
Thyroxine 20
UTP 20
PRPP 50
Acetylcholine 25
Adenosine-5'-phosphate 5
Cholesterol 5
Coenzyme A 50
Deoxycytidine HCl 15
Deoxyribose 50
DPN 10
Fructose-1-6-diphosphate
120
Glucosamine 50
Glucuronate 25
Glutathione 20
Guanine 50
D-Ribose 5
Sodium pyruvate 150
Thymidine 25
TPN 20
Xanthine 25
Glycogen 250
Calcium Chloride 150
Potassium Chloride 500
Magnesium Sulfate 250
Ferric Ammonium Sulfate
20
KH2PO4 75
Sodium Chloride 6500
Sodium Hydrogen Phosphate, disodium
15
Magnesium Chloride 500
L-Lecithin 150
and Sphingomyelin 75,
______________________________________ and Sodium bicarbonate--upto 1500 mg/liter, sufficient to adjust final pH to 7.4.
Claim 2
Dependent Claim
273 words
(a) A cell culturing medium comprising an amino acid composition containing: ______________________________________
Item mg/liter
______________________________________
Alanine 60
Alpha-Amino-n-butyric Acid
5
Arginine 150
Asparagine 5
Aspartic Acid 75
Cysteine-HCl 100
Cystine 25
Ergothionine 10
Glutamic Acid 500
Glutamine 800
Glycine 500
Histidine, HCl 50
Homocysteine 20
Isoleucine 75
Leucine 300
Lysine HCl 300
Methionine 50
Ornithine HCl 10
Phenylalanine 100
Proline 100
Serine 75
Taurine 50
Threonine 100
Tryptophane 50
Tyrosine 100
and Valine 200,
______________________________________ (b) a vitamin mixture containing: ______________________________________
Item mg/liter
______________________________________
DL-Alpha-Tocopherol
2.5
Ascorbic Acid 25
Biotin 20
Calciferol 2.5
Calcium-pentothenate
25
Choline Chloride 25
Cocarboxylase 20
Folic Acid 20
i-Inositol 15
Menadione 2.5
Niacin 5.0
Niacinamide 20
p-Aminobenzoic Acid
1.0
Pyridoxal HCl 20
Pyridoxine HCl 5.0
Riboflavin 20
Thiamine HCl 20
Vitamin B12 50
Vitamin A 10
and Citric Acid 20,
______________________________________ (c) other components containing: ______________________________________
Item mg/liter
______________________________________
Na--Acetate, 3 H.sub.2 O
100
ATP (Sodium Salt) 20
Estradiol 20
Cholate 25
Deoxyadenosine 15
Deoxyguanosine 15
Dextrose 1500
Flavin Adenine Dinucleotide
20
Galactosamine 120
Glucose-1-phosphate 25
Glucuronolactone 25
Hypoxanthine 100
5-Methyl-deoxycytidylic Acid
5
Phenol Red 15
Thioactic Acid 2
Thyroxine 20
UTP 20
PRPP 50
Acetylcholine 25
Adenosine-5'-phosphate 5
Cholesterol 5
Coenzyme A 50
Deoxycytidine HCl 15
Deoxyribose 50
DPN 10
Fructose-1-6-diphosphate
120
Glucosamine 50
Glucuronate 25
Glutathione 20
Guanine 50
D-Ribose 5
Sodium pyruvate 150
Thymidine 25
TPN 20
Xanthine 25
Glycogen 250
Calcium Chloride 150
Potassium Chloride 500
Magnesium Sulfate 250
Ferric Ammonium Sulfate
20
KH2PO4 75
Sodium Chloride 6500
Sodium Hydrogen Phosphate, disodium
15
Magnesium Chloride 500
L-Lecithin 150
and Sphingomyelin 75,
______________________________________ and Sodium bicarbonate--upto 1500 mg/liter, sufficient to adjust final pH to 7.4.
Claim 3
Dependent Claim
121 words
A culture medium as defined in claim 1, in which the concentrations of the vitamin mixture identified in subsection (b) are in the following range: ______________________________________
Item mg/liter
______________________________________
DL-Alpha-Tocopherol
2.0 to 3.0
Ascorbic Acid 20.0 to 30.0
Biotin 10.0 to 25.0
Calciferol 1.0 to 2.8
Calcium-pentothenate
15.0 to 25.0
Choline Chloride 15.0 to 25.0
Cocarboxylase 10.0 to 20.0
Folic Acid 10.0 to 20.0
i-Inositol 10.0 to 20.0
Menadione 2.0 to 3.0
Niacin 4.0 to 5.0
Niacinamide 4.0 to 20.0
p-Aminobenzoic Acid
0.1 to 1.0
Pyridoxal HCl 5.0 to 20.0
Pyridoxine HCl 2.0 to 5.0
Riboflavin 5.0 to 35.0
Thiamine HCl 5.0 to 35.0
Vitamin B12 10.0 to 50.0
Vitamin A 5.0 to 20.0
and Citric Acid 10.0 to 20.0
______________________________________
Claim 4
Dependent Claim
13 words
A culture media, as defined in claim 2, further comprising: Gentamycin: 5.0 mg/liter.
Claim 5
Dependent Claim
25 words
A culture media, as defined in claim 3, further comprising: ______________________________________
Item mg/liter
______________________________________
Colchicine 0.0025
Colcemid 0.0025
EDTA 0.0050
and Phytohemagglutinin 5.0 ml/liter
______________________________________
Claim 6
Dependent Claim
13 words
A culture media, as defined in claim 1, further comprising: Gentamycin: 5.0 mg/liter.
Claim 7
Dependent Claim
25 words
A culture media, as defined in claim 3, further comprising: ______________________________________
Item mg/liter
______________________________________
Colchicine 0.0025
Colcemid 0.0025
EDTA 0.0050
and Phytohemagglutinin 5.0 ml/liter
______________________________________
Claim 8
Dependent Claim
262 words
An aqueous culture media, for use in cell growth comprising: ______________________________________
Item mg/liter
______________________________________
Alanine 60
Alpha-Amino-n-butyric Acid
5
Arginine 150
Asparagine 5
Aspartic Acid 75
Cysteine-HCl 100
Cystine 25
Ergothionine 10
Glutamic Acid 500
Glutamine 800
Glycine 500
Histidine, HCl 50
Homocysteine 20
Isoleucine 75
Leucine 300
Lysine HCl 300
Methionine 50
Ornithine HCl 10
Phenylalanine 100
Proline 100
Serine 75
Taurine 50
Threonine 100
Tryptophane 50
Tyrosine 100
Valine 200
DL-Alpha-Tocopherol 2.5
Ascorbic Acid 25
Biotin 20
Calciferol 2.5
Calcium-pentothenate 25
Choline Chloride 25
Cocarboxylase 20
Folic Acid 20
i-Inositol 15
Menadione 2.5
Niacin 5.0
Niacinamide 20
p-Aminobenzoic Acid 1.0
Pyridoxal HCl 20
Pyridoxine HCl 5.0
Riboflavin 20
Thiamine HCl 20
Vitamin B12 50
Vitamin A 10
Citric Acid 20
Na--Acetate, 3 H2O 100
ATP (Sodium salt) 20
Estradiol 20
Cholate 25
Deoxyadenosine 15
Deoxyguanosine 15
Dextrose 1500
Flavin Adenine Dinucleotide
20
Galactosamine 120
Glucose-1-phosphate 25
Glucuronolactone 25
Hypoxanthine 100
5-Methyl-deoxycytidylic Acid
5
Phenol Red 15
Thioactic Acid 2
Thyroxine 20
UTP 20
PRPP 50
Acetylcholine 25
Adenosine-5'-phosphate 5
Cholesterol 5
Coenzyme A 50
Deoxycytidine HCl 15
Deoxyribose 50
DPN 10
Fructose-1-6-diphosphate
120
Glucosamine 50
Glucuronate 25
Glutathione 20
Guanine 50
D-Ribose 5
Sodium pyruvate 150
Thymidine 25
TPN 20
Xanthine 25
Glycogen 250
Calcium Chloride 150
Potassium Chloride 500
Magnesium Sulfate 250
Ferric Ammonium Sulfate 20
KH2PO4 75
Sodium Chloride 6500
Sodium Hydrogen Phosphate, disodium
15
Magnesium Chloride 500
L-Lecithin 150
Sphingomyelin 75
______________________________________
Sodium bicarbonate up to 1500 mg/liter, sufficient to adjust final pH to
7.4 and Sodium bicarbonate--upto 1500 mg/liter, sufficient to adjust final pH to 7.4.
Claim 9
Dependent Claim
262 words
An aqueous culture media, for use in cell growth comprising: ______________________________________
Item mg/liter
______________________________________
Alanine 60
Alpha-Amino-n-butyric Acid
5
Arginine 150
Asparagine 5
Aspartic Acid 75
Cysteine-HCl 100
Cystine 25
Ergothionine 10
Glutamic Acid 500
Glutamine 800
Glycine 500
Histidine, HCl 50
Homocysteine 20
Isoleucine 75
Leucine 300
Lysine HCl 300
Methionine 50
Ornithine HCl 10
Phenylalanine 100
Proline 100
Serine 75
Taurine 50
Threonine 100
Tryptophane 50
Tyrosine 100
Valine 200
DL-Alpha-Tocopherol 2.5
Ascorbic Acid 25
Biotin 20
Calciferol 2.5
Calcium-pentothenate 25
Choline Chloride 25
Cocarboxylase 20
Folic Acid 20
i-Inositol 15
Menadione 2.5
Niacin 5.0
Niacinamide 20
p-Aminobenzoic Acid 1.0
Pyridoxal HCl 20
Pyridoxine HCl 5.0
Riboflavin 20
Thiamine HCl 20
Vitamin B12 50
Vitamin A 10
Citric Acid 20
Na--Acetate, 3 H2O 100
ATP (Sodium salt) 20
Estradiol 20
Cholate 25
Deoxyadenosine 15
Deoxyguanosine 15
Dextrose 1500
Flavin Adenine Dinucleotide
20
Galactosamine 120
Glucose-1-phosphate 25
Glucuronolactone 25
Hypoxanthine 100
5-Methyl-deoxycytidylic Acid
5
Phenol Red 15
Thioactic Acid 2
Thyroxine 20
UTP 20
PRPP 50
Acetylcholine 25
Adenosine-5'-phosphate 5
Cholesterol 5
Coenzyme A 50
Deoxycytidine HCl 15
Deoxyribose 50
DPN 10
Fructose-1-6-diphosphate
120
Glucosamine 50
Glucuronate 25
Glutathione 20
Guanine 50
D-Ribose 5
Sodium pyruvate 150
Thymidine 25
TPN 20
Xanthine 25
Glycogen 250
Calcium Chloride 150
Potassium Chloride 500
Magnesium Sulfate 250
Ferric Ammonium Sulfate 20
KH2PO4 75
Sodium Chloride 6500
Sodium Hydrogen Phosphate, disodium
15
Magnesium Chloride 500
L-Lecithin 150
Sphingomyelin 75
______________________________________
Sodium bicarbonate up to 1500 mg/liter, sufficient to adjust final pH to
7.4 and Sodium bicarbonate--upto 1500 mg/liter, sufficient to adjust final pH to 7.4.
Claim 10
Dependent Claim
112 words
An aqueous culture media as defined in claim 7, wherein the vitamin concentrations are as follows: ______________________________________
Item mg/liter
______________________________________
DL-Alpha-Tocopherol
2.0 to 3.0
Ascorbic Acid 20.0 to 30.0
Biotin 10.0 to 25.0
Calciferol 1.0 to 2.8
Calcium-pentothenate
15.0 to 25.0
Choline Chloride 15.0 to 25.0
Cocarboxylase 10.0 to 20.0
Folic Acid 10.0 to 20.0
i-Inositol 10.0 to 20.0
Menadione 2.0 to 3.0
Niacin 4.0 to 5.0
Niacinamide 4.0 to 20.0
p-Aminobenzoic Acid
0.1 to 1.0
Pyridoxal HCl 5.0 to 20.0
Pyridoxine HCl 2.0 to 5.0
Riboflavin 5.0 to 35.0
Thamie HCl 5.0 to 35.0
Vitamin B12 10.0 to 50.0
Vitamin A 5.0 to 20.0
and Citric Acid 10.0 to 20.0.
______________________________________
Claim 11
Dependent Claim
25 words
A culture media, as defined in any one of the claims 1 thru 8, wherein said medium is free of cell-damaging qualities of proteolytic enzymes.
Claim 12
Independent Claim
28 words
A method for growing human amniotic fluid cells comprising culturing human amniotic fluid cells in the culture media as defined in any one of claims 1 to 8.
Claim 13
Independent Claim
28 words
A method for growing human or animal neural tissues comprising culturing human or animal neural cells in the culture media defined in anyone of claims 1 to 8.
References Cited (Backward Citations)
No citation records indexed.
USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 1990-04-13 | 1990-10-13 | 1991-04-14 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1994-04-13 | 1994-10-13 | 1995-04-14 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 1998-04-13 | 1998-10-13 | 1999-04-14 | $7700.00 | Window Closed / Lapsed |