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US5405772A
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Medium for long-term proliferation and development of cells
Assignee: Amgen Inc
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Filing: 1993-06-18
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Granted: 1995-04-11
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Expires: 2013-06-18
Abstract
Abstract A serum-free or serum-depleted medium for the short- and long-term proliferation and development of cells, particularly hematopoietic cells and bone marrow stromal cells, the medium comprising cell proliferation and development effective amounts of: a standard culture medium such as Iscove's modified Dulbecco's medium; serum albumin; transferrin; a source of lipids and fatty acids; cholesterol; a reducing agent; pyruvate; a glucocorticoid (when the cells to be cultured are hematopoietic cells); nucleosides for synthesis of DNA and RNA; growth factors that stimulate the proliferation and development of stromal cells and cells from a variety of tissues or organs, such as epidermal growth factor, fibroblast growth factor, platelet derived growth factor, and insulin; and extracellular matrix materials, such as collagen, fibronectin, and laminin.
Inventors
- 👤 Ian L. O. Ponting
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-11-03 22:19
CPC Classifications
C; C12; C12N; C12N5/00; C12N5/06; C12N5/0602; C12N5/0652; C12N5/0662; C12N5/0663; C12N5/0634; C12N5/0647; C12N2500/00; C12N2500/05; C12N2500/10; C12N2500/20; C12N2500/24; C12N2500/25; C12N2500/30; C12N2500/36; C12N2500/40; C12N2500/90; C12N2501/00; C12N2501/10; C12N2501/11; C12N2501/115; C12N2501/135; C12N2501/30; C12N2501/38; C12N2501/39; C12N2501/50; C12N2501/58; C12N2502/00; C12N2502/13; C12N2502/1394
Claims Analysis
Total Claims: 17
Claim 1
Independent Claim
156 words
A medium for the long-term proliferation and development of cells, which comprises the following ingredients in the indicated amounts: ______________________________________
(a) a standard culture medium
0.8-1.09 x,
(b) serum albumin 3-50 mg/ml,
(c) transferrin 25-1000 ug/ml,
(d) lipids and fatty acids
5-100 ug/ml,
(e) cholesterol 3-30 ug/ml,
(f) reducing agent 30-300 uM,
(g) pyruvate 30-500 ug/ml,
(h) nucleosides 5-30 ug/ml,
______________________________________ (i) at least one growth factor selected from the group consisting of: ______________________________________
(1) epidermal growth factor
5-200 ng/ml
(2) fibroblast growth factor
0.5-40 ng/ml
(3) platelet-derived growth factor
2-200 ng/ml; and
(4) insulin 2-100 ug/ml.
______________________________________ (j) at least one extracellular matrix material selected from the group consisting of: ______________________________________
(1) collagen IV
2-100 ug/cm.sup.2 ; and
(2) fibronectin
0.5-100 ug/cm.sup.2 ;
______________________________________ wherein said medium for the long-term proliferation and development of cells is serum-free or serum-depleted and is capable of culturing cells comprising adipocytes, macrophages, endothelial cells, fibroblasts, and hematopoictic progenitor cells.
Claim 2
Independent Claim
156 words
A medium for the long-term proliferation and development of cells, which comprises the following ingredients in the indicated amounts: ______________________________________
(a) a standard culture medium
0.8-1.09 x,
(b) serum albumin 3-50 mg/ml,
(c) transferrin 25-1000 ug/ml,
(d) lipids and fatty acids
5-100 ug/ml,
(e) cholesterol 3-30 ug/ml,
(f) reducing agent 30-300 uM,
(g) pyruvate 30-500 ug/ml,
(h) nucleosides 5-30 ug/ml,
______________________________________ (i) at least one growth factor selected from the group consisting of: ______________________________________
(1) epidermal growth factor
5-200 ng/ml
(2) fibroblast growth factor
0.5-40 ng/ml
(3) platelet-derived growth factor
2-200 ng/ml; and
(4) insulin 2-100 ug/ml.
______________________________________ (j) at least one extracellular matrix material selected from the group consisting of: ______________________________________
(1) collagen IV
2-100 ug/cm.sup.2 ; and
(2) fibronectin
0.5-100 ug/cm.sup.2 ;
______________________________________ wherein said medium for the long-term proliferation and development of cells is serum-free or serum-depleted and is capable of culturing cells comprising adipocytes, macrophages, endothelial cells, fibroblasts, and hematopoictic progenitor cells.
Claim 3
Dependent Claim
10 words
The medium according to claim 1, further comprising a glucocorticoid.
Claim 4
Dependent Claim
11 words
The medium according to claim 1, wherein said glucocorticoid is hydrocortisone.
Claim 5
Dependent Claim
10 words
The medium according to claim 1, further comprising stromal cells.
Claim 6
Dependent Claim
13 words
The medium according to claim 1, further comprising an inhibitor of CSF-1 activity.
Claim 7
Dependent Claim
17 words
The medium according to claim 1, wherein said cells to be cultured are incapable of producing CSF-1.
Claim 8
Dependent Claim
19 words
The medium according to claim 6, wherein said cells to be cultured are obtained from a B6C3Fe-a/a-op/op mouse strain.
Claim 9
Dependent Claim
16 words
The medium according to claim 1, wherein said standard culture medium is Iscove's Modified Dulbecco's Medium.
Claim 10
Dependent Claim
22 words
The medium according to claim 1, wherein said serum albumin, transferrin and growth factors are each of human, mouse or bovine origin.
Claim 11
Dependent Claim
28 words
The medium according to claim 1, wherein said nucleosides are one or more selected from the group consisting of adenosine, guanosine, cytidine, uridine, 2'-deoxyadenosine, 2'-deoxyguanosine, 2'-deoxycytidine and thymidine.
Claim 12
Dependent Claim
15 words
The medium according to claim 1, wherein all of said extracellular matrix materials are included.
Claim 13
Dependent Claim
13 words
The medium according to claim 1, wherein said cells are hematopoietic or lymphopoietic.
Claim 14
Dependent Claim
15 words
The medium according to claim 1, wherein said cells are from mammalian organs or tissues.
Claim 15
Dependent Claim
15 words
The medium according to claim 1, wherein said cells are primary cells or cell lines.
Claim 16
Dependent Claim
24 words
A method of long-term culture of cells which comprises culturing the cells for 8 to 20 weeks in the medium according to claim 1.
Claim 17
Dependent Claim
24 words
A method of short-term culture of cells which comprises culturing the cells for 2 to 7 weeks in the medium according to claim 1.
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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) | 1998-04-10 | 1998-10-10 | 1999-04-11 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 2002-04-10 | 2002-10-10 | 2003-04-11 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 2006-04-10 | 2006-10-10 | 2007-04-11 | $7700.00 | Window Closed / Lapsed |