US7439064B2
Expired / Lapsed
US
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Cultivation of human embryonic stem cells in the absence of feeder cells or without conditioned medium
Assignee: Wicell Research Institute Inc
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Filing: 2005-03-11
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Granted: 2008-10-21
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Expires: 2025-03-11
Abstract
Abstract The invention relates to methods for culturing human embryonic stem cells by culturing the stem cells in an environment essentially free of mammalian fetal serum and in a stem cell culture medium including amino acids, vitamins, salts, minerals, transferring, insulin, albumin, and a fibroblast growth factor that is supplied from a source other than just a feeder layer the medium. Also disclosed are compositions capable of supporting the culture and proliferation of human embryonic stem cells without the need for feeder cells or for exposure of the medium to feeder cells.
Inventors
- 👤 James A. Thomson
- 👤 Mark Levenstein
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-11-03 00:45
CPC Classifications
C; C12; C12N; C12N5/00; C12N5/06; C12N5/0602; C12N5/0603; C12N5/0606; C12N2500/00; C12N2500/05; C12N2500/10; C12N2500/20; C12N2500/24; C12N2500/25; C12N2500/30; C12N2500/44; C12N2500/90; C12N2501/00; C12N2501/10; C12N2501/115
Claims Analysis
Total Claims: 14
Claim 1
Independent Claim
105 words
A method of culturing human embryonic stem cells, comprising: culturing human embryonic stem cells on a matrix in a culture medium free of serum and in a stem cell culture medium containing amino acids, vitamins, salts, minerals, transferrin or a transferrin substitute, insulin or an insulin substitute, albumin, and a fibroblast growth factor supplied from a source other than a feeder layer, the fibroblast growth factor present in a concentration at least as high as a maintenance concentration, wherein the medium supports the culture and proliferation of undifferentiated proliferating euploid human embryonic stem cells for at least six passages without feeder cells or conditioned medium.
Claim 2
Independent Claim
105 words
A method of culturing human embryonic stem cells, comprising: culturing human embryonic stem cells on a matrix in a culture medium free of serum and in a stem cell culture medium containing amino acids, vitamins, salts, minerals, transferrin or a transferrin substitute, insulin or an insulin substitute, albumin, and a fibroblast growth factor supplied from a source other than a feeder layer, the fibroblast growth factor present in a concentration at least as high as a maintenance concentration, wherein the medium supports the culture and proliferation of undifferentiated proliferating euploid human embryonic stem cells for at least six passages without feeder cells or conditioned medium.
Claim 3
Dependent Claim
17 words
The method of claim 1 wherein the FGF is selected from FGF2, FGF4, FGF9, FGF17 and FGF18.
Claim 4
Dependent Claim
19 words
The method of claim 1 wherein the FGF is FGF2 which is present in the medium at 100 ng/ml.
Claim 5
Independent Claim
93 words
A method of culturing human embryonic stem cells in defined media without serum and without feeder cells on a matrix, the method comprising: culturing human embryonic stem cells in a culture medium containing albumin, amino acids, vitamins, minerals, at least one transferrin or transferrin substitute, at least one insulin or insulin substitute, the culture medium free of serum and containing at least about 100 ng/ml of a fibroblast growth factor, and culturing without feeder cells or conditioned medium, wherein the medium supports proliferation of the human embryonic stem cells in an undifferentiated state.
Claim 6
Independent Claim
93 words
A method of culturing human embryonic stem cells in defined media without serum and without feeder cells on a matrix, the method comprising: culturing human embryonic stem cells in a culture medium containing albumin, amino acids, vitamins, minerals, at least one transferrin or transferrin substitute, at least one insulin or insulin substitute, the culture medium free of serum and containing at least about 100 ng/ml of a fibroblast growth factor, and culturing without feeder cells or conditioned medium, wherein the medium supports proliferation of the human embryonic stem cells in an undifferentiated state.
Claim 7
Dependent Claim
51 words
The method of claim 4 , wherein said culturing step includes the embryonic stem cells proliferating in culture for over one month while maintaining the potential of the embryonic stem cells to differentiate into derivatives of endoderm, mesoderm, and ectoderm tissues, and while maintaining the karyotype of the embryonic stem cells.
Claim 8
Dependent Claim
17 words
The method of claim 4 wherein the FGF is selected from FGF2, FGF4, FGF9, FGF17 and FGF18.
Claim 9
Dependent Claim
127 words
A culture of human embryonic stem cells comprising: human embryonic stem cells, a matrix; and a stem cell medium containing albumin, amino acids, vitamins, minerals, at least one transferrin or transferrin substitute, at least one insulin or insulin substitute, the culture medium free of serum and containing a fibroblast growth factor supplied from a source other than a feeder layer, the fibroblast growth factor present in a concentration at least as high as a maintenance concentration, wherein the medium supports the culture of the embryonic stem cells indefinitely in the absence of serum and in the absence of feeder cells and also in the absence of medium exposed to feeder cells, wherein the culture maintains the embryonic stem cells in an undifferentiated state indefinitely with normal karyotype.
Claim 10
Dependent Claim
127 words
A culture of human embryonic stem cells comprising: human embryonic stem cells, a matrix; and a stem cell medium containing albumin, amino acids, vitamins, minerals, at least one transferrin or transferrin substitute, at least one insulin or insulin substitute, the culture medium free of serum and containing a fibroblast growth factor supplied from a source other than a feeder layer, the fibroblast growth factor present in a concentration at least as high as a maintenance concentration, wherein the medium supports the culture of the embryonic stem cells indefinitely in the absence of serum and in the absence of feeder cells and also in the absence of medium exposed to feeder cells, wherein the culture maintains the embryonic stem cells in an undifferentiated state indefinitely with normal karyotype.
Claim 11
Dependent Claim
27 words
The culture of claim 7 wherein the fibroblast growth factor is FGF2 which is present in the medium in a concentration of at least about 100 ng/ml.
Claim 12
Dependent Claim
107 words
A culture of feeder independent human embryonic stem cells comprising human embryonic stem cells on a matrix in a stem cell culture medium, the stem cell culture medium comprising albumin, amino acids, vitamins, minerals, at least one transferrin or transferrin substitute, at least one insulin or insulin substitute, the culture medium free of serum and containing a fibroblast growth factor present in a concentration at least as high as a maintenance concentration, wherein the fibroblast growth factor is selected from FGF2, FGF4, FGF9, FGF17, and FGF18, wherein the culture is independent of feeder cells while the human embryonic stem cells remain euploid and in an undifferentiated state.
Claim 13
Dependent Claim
107 words
A culture of feeder independent human embryonic stem cells comprising human embryonic stem cells on a matrix in a stem cell culture medium, the stem cell culture medium comprising albumin, amino acids, vitamins, minerals, at least one transferrin or transferrin substitute, at least one insulin or insulin substitute, the culture medium free of serum and containing a fibroblast growth factor present in a concentration at least as high as a maintenance concentration, wherein the fibroblast growth factor is selected from FGF2, FGF4, FGF9, FGF17, and FGF18, wherein the culture is independent of feeder cells while the human embryonic stem cells remain euploid and in an undifferentiated state.
Claim 14
Dependent Claim
23 words
A culture as claimed in claim 9 wherein the fibroblast growth factor is present at a concentration of at least about 100 ng/ml.
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USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 2011-10-21 | 2012-04-21 | 2012-10-21 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 2015-10-21 | 2016-04-21 | 2016-10-21 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 2019-10-21 | 2020-04-21 | 2020-10-21 | $7700.00 | Window Closed / Lapsed |