US5114855A
Expired / Lapsed
US
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Method for aggregating cells with small microspheres
Assignee: University of Minnesota System
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Filing: 1990-04-19
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Granted: 1992-05-19
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Expires: 2010-04-19
Abstract
Abstract A method of inducing aggregate formation of animal cells is carried out by forming a nutrient medium suspension of animal cells and introducing itno the suspension microspheres of a derivatized diameter of not more than about 60 μm, when measured in a suitable buffer saline or cell culture media.
Inventors
- 👤 Wei-Shou Hu
- 👤 Stephane Goetghebeur
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-10-23 04:34
CPC Classifications
C; C12; C12N; C12N5/00; C12N5/0068; C12N5/0075; C12M; C12M25/00; C12M25/16; C12N2533/00; C12N2533/70
Claims Analysis
Total Claims: 55
Claim 1
Independent Claim
52 words
A method of inducing aggregate formation of anchorage-dependent cells comprising the steps of: (a) forming a suspension of anchorage-dependent cells in nutrient medium; and (b) introducing into the suspension microspheres having a positive finite derivatized diameter of less than about 60 μm, when measured in a buffer saline or cell culture media.
Claim 2
Independent Claim
52 words
A method of inducing aggregate formation of anchorage-dependent cells comprising the steps of: (a) forming a suspension of anchorage-dependent cells in nutrient medium; and (b) introducing into the suspension microspheres having a positive finite derivatized diameter of less than about 60 μm, when measured in a buffer saline or cell culture media.
Claim 3
Independent Claim
65 words
A method of inducing aggregate formation of cells which grow in suspension or on surface comprising the steps of: (a) forming a uniform nutrient medium dispersion of cells which grow in suspension or on surface; and (b) introducing into the dispersion microspheres having a positive finite derivatized diameter of less than about 60 μm, when measured in a suitable buffer saline or cell culture media.
Claim 4
Independent Claim
65 words
A method of inducing aggregate formation of cells which grow in suspension or on surface comprising the steps of: (a) forming a uniform nutrient medium dispersion of cells which grow in suspension or on surface; and (b) introducing into the dispersion microspheres having a positive finite derivatized diameter of less than about 60 μm, when measured in a suitable buffer saline or cell culture media.
Claim 5
Dependent Claim
50 words
In a method of inducing aggregate formation of anchorage-dependent cells in a nutrient medium suspension of said anchorage-dependent cells, the improvement comprising introducing into said suspension microspheres of a positive finite derivatized diameter of less than about 60 μm, when measured in a suitable buffer saline or cell culture media.
Claim 6
Dependent Claim
50 words
In a method of inducing aggregate formation of anchorage-dependent cells in a nutrient medium suspension of said anchorage-dependent cells, the improvement comprising introducing into said suspension microspheres of a positive finite derivatized diameter of less than about 60 μm, when measured in a suitable buffer saline or cell culture media.
Claim 7
Dependent Claim
55 words
In a method of inducing aggregate formation of suspension-grown or surface-grown cells in a uniform nutrient medium dispersion of said suspension-grown or surface-grown cells, the improvement comprising introducing into said dispersion microspheres of a positive finite derivatized diameter of less than about 60 μm, when measured in a suitable buffer saline or cell culture media.
Claim 8
Dependent Claim
55 words
In a method of inducing aggregate formation of suspension-grown or surface-grown cells in a uniform nutrient medium dispersion of said suspension-grown or surface-grown cells, the improvement comprising introducing into said dispersion microspheres of a positive finite derivatized diameter of less than about 60 μm, when measured in a suitable buffer saline or cell culture media.
Claim 9
Dependent Claim
19 words
A method according to claim 1, wherein the microspheres are of a diameter of approximately 15 to 25 μm.
Claim 10
Dependent Claim
19 words
A method according to claim 2, wherein the microspheres are of a diameter of approximately 15 to 25 μm.
Claim 11
Dependent Claim
19 words
The improvement according to claim 3, wherein the microspheres are of a diameter of approximately 15 to 25 μm.
Claim 12
Dependent Claim
19 words
The improvement according to claim 4, wherein the microspheres are of a diameter of approximately 15 to 25 μm.
Claim 13
Dependent Claim
21 words
A method according to claim 2, wherein the cells are selected from Chinese hamster ovary cells or human kidney cells 293.
Claim 14
Dependent Claim
20 words
A method according to claim 1, wherein the cells are selected from swine testicular cells or monkey kidney cells, Vero.
Claim 15
Dependent Claim
20 words
The improvement according to claim 3, wherein the cells are selected from swine testicular cells or monkey kidney cells, Vero.
Claim 16
Dependent Claim
21 words
The improvement according to claim 4, wherein the cells are selected from Chinese hamster ovary cells or human kidney cells 293.
Claim 17
Dependent Claim
24 words
A method according to claim 1, wherein the microspheres are able to accommodate a monolayer cell density of 10 to 20 cells per microsphere.
Claim 18
Dependent Claim
24 words
A method according to claim 2, wherein the microspheres are able to accommodate a monolayer cell density of 10 to 20 cells per microsphere.
Claim 19
Dependent Claim
24 words
The improvement according to claim 3, wherein the microspheres are able to accommodate a monolayer cell density of 10 to 20 cells per microsphere.
Claim 20
Dependent Claim
21 words
A method according to claim 1, wherein the microsphere concentration in the suspension formed by step (b) is approximately 0.5 g/l.
Claim 21
Dependent Claim
21 words
A method according to claim 2, wherein the microsphere concentration in the suspension formed by step (b) is approximately 0.5 g/l.
Claim 22
Dependent Claim
21 words
The improvement according to claim 3, wherein the microsphere concentration in the suspension formed by step (b) is approximately 0.5 g/l.
Claim 23
Dependent Claim
21 words
The improvement according to claim 4, wherein the microsphere concentration in the suspension formed by step (b) is approximately 0.5 g/l.
Claim 24
Independent Claim
54 words
A method of inducing aggregate formation of animal cells comprising the steps of: (a) forming a suspension of animal cells in nutrient medium; and (b) introducing into the suspension microspheres of a positive finite derivatized diameter of not more than about 60 μm, when measured in a suitable buffer saline or cell culture media.
Claim 25
Independent Claim
54 words
A method of inducing aggregate formation of animal cells comprising the steps of: (a) forming a suspension of animal cells in nutrient medium; and (b) introducing into the suspension microspheres of a positive finite derivatized diameter of not more than about 60 μm, when measured in a suitable buffer saline or cell culture media.
Claim 26
Dependent Claim
51 words
In a method of inducing aggregate formation of animal cells in a nutrient medium suspension of said animal cells, the improvement comprising introducing into said suspension microspheres of a positive finite derivatized diameter of not more than about 60 μm, when measured in a suitable buffer saline or cell culture media.
Claim 27
Dependent Claim
51 words
In a method of inducing aggregate formation of animal cells in a nutrient medium suspension of said animal cells, the improvement comprising introducing into said suspension microspheres of a positive finite derivatized diameter of not more than about 60 μm, when measured in a suitable buffer saline or cell culture media.
Claim 28
Independent Claim
44 words
A method of inducing aggregate formation of animal cells by introducing into a nutrient medium suspension of animal cells microspheres of a positive finite derivatized diameter of not more than about 60 μm, when measured in a suitable buffer saline or cell culture media.
Claim 29
Independent Claim
44 words
A method of inducing aggregate formation of animal cells by introducing into a nutrient medium suspension of animal cells microspheres of a positive finite derivatized diameter of not more than about 60 μm, when measured in a suitable buffer saline or cell culture media.
Claim 30
Dependent Claim
48 words
In a method of inducing aggregate formation of animal cells, the improvement comprising introducing into a suspension of animal cell sin nutrient medium microspheres of a positive finite derivatized diameter of not more than about 60 μm, when measured in a suitable buffer saline or cell culture media.
Claim 31
Dependent Claim
48 words
In a method of inducing aggregate formation of animal cells, the improvement comprising introducing into a suspension of animal cell sin nutrient medium microspheres of a positive finite derivatized diameter of not more than about 60 μm, when measured in a suitable buffer saline or cell culture media.
Claim 32
Dependent Claim
26 words
A method according to claim 1, wherein the microspheres are formed of dextran, gelatin, polyacrylamide-copolymerized with collagen or gelatin, polyacrylamide with modified charge, polystyrene or glass.
Claim 33
Dependent Claim
26 words
A method according to claim 2, wherein the microspheres are formed of dextran, gelatin, polyacrylamide-copolymerized with collagen or gelatin, polyacrylamide with modified charge, polystyrene or glass.
Claim 34
Dependent Claim
26 words
The improvement according to claim 3, wherein the microspheres are formed of dextran, gelatin, polyacrylamide-copolymerized with collagen or gelatin, polyacrylamide with modified charge, polystyrene or glass.
Claim 35
Dependent Claim
26 words
The improvement according to claim 4, wherein the microspheres are formed of dextran, gelatin, polyacrylamide-copolymerized with collagen or gelatin, polyacrylamide with modified charge, polystyrene or glass.
Claim 36
Dependent Claim
49 words
In a method of growing anchorage dependent cells in suspension, the improvement comprising employing in the suspension as a microcarrier support for attachment of cells, microspheres of a positive finite derivatized diameter of less than about 60 μm, when measured in a suitable buffer saline or cell culture media.
Claim 37
Dependent Claim
49 words
In a method of growing anchorage dependent cells in suspension, the improvement comprising employing in the suspension as a microcarrier support for attachment of cells, microspheres of a positive finite derivatized diameter of less than about 60 μm, when measured in a suitable buffer saline or cell culture media.
Claim 38
Dependent Claim
54 words
A method according to claim 1, in which the microsphere diameter is chosen so that a mean of cells/microsphere at initiation of culture is at least approximately 4 to 5 cells/microsphere and so that rate of cell attachment to microsphere surface at initiation of culture exhibits zero order kinetics with respect to surface area.
Claim 39
Dependent Claim
52 words
A method according to claim 2, in which the microsphere diameter is chosen sot hat at initiation of culture a mean of cells/microsphere is approximately 4 to 5 cells/microsphere and so that at initiation of culture rate of cell attachment to microsphere surface exhibits zero order kinetics with respect to surface area.
Claim 40
Dependent Claim
54 words
A method according to claim 3, in which the diameter of the microsphere is chosen so that a mean of cells/microsphere at initiation of culture is approximately 4 to 5 cells/microsphere and so that rate of cell attachment to microsphere surface at imitation of culture exhibits zero order kinetics with respect to surface area.
Claim 41
Dependent Claim
54 words
A method according to claim 4, in which the diameter of the microsphere is chosen sot at a mean of cells/microsphere at initiation of culture is approximately 4 to 5 cells/microsphere and so that rate of cell attachment to microsphere surface at initiation of culture exhibits zero order kinetics with respect to surface area.
Claim 42
Dependent Claim
54 words
A method according to claim 20, in which the diameter of the microsphere is chosen sot hat a mean of cells/microsphere at initiation of culture is approximately 4 to 5 cells/microsphere and so that rate of cell attachment to microsphere surface at initiation of culture exhibits zero order kinetics with respect to surface area.
Claim 43
Dependent Claim
54 words
A method according to claim 21, in which the diameter of the microsphere is chosen so that a mean of cells/microsphere at initiation of culture is approximately 4 to 5 cells/microsphere and so that rate of cell attachment to microsphere surface at initiation of culture exhibits zero order kinetics with respect to surface area.
Claim 44
Dependent Claim
54 words
A method according to claim 22, in which the diameter of the microsphere is chosen so that a mean of cells/microsphere at initiation of culture is approximately 4 to 5 cells/microsphere and so that rate of cell attachment to microsphere surface at initiation of culture exhibits zero order kinetics with respect to surface area.
Claim 45
Dependent Claim
50 words
A method according to claim 23, in which the diameter of the microsphere is chosen so that a mean of cells/microsphere at initiation of culture is approximately 4 to 5 cells/microsphere and so that rate of cell attachment to microsphere surface exhibits zero order kinetics with respect to surface area.
Claim 46
Dependent Claim
54 words
A method according to claim 28, in which the diameter of the microsphere is chosen so that a mean of cells/microsphere at initiation of culture is approximately 4 to 5 cells/microsphere and so that rate of cell attachment to microsphere surface at initiation of culture exhibits zero order kinetics with respect to surface area.
Claim 47
Dependent Claim
19 words
A method according to claim 1, wherein the diameter of the microspheres is a minimum of about 10 μm.
Claim 48
Dependent Claim
19 words
A method according to claim 2, wherein the diameter of the microspheres is a minimum of about 10 μm.
Claim 49
Dependent Claim
19 words
A method according to claim 3, wherein the diameter of the microspheres is a minimum of about 10 μm.
Claim 50
Dependent Claim
19 words
A method according to claim 4, wherein the diameter of the microspheres is a minimum of about 10 μm.
Claim 51
Dependent Claim
19 words
A method according to claim 20, wherein the diameter of the microspheres is a minimum of about 10 μm.
Claim 52
Dependent Claim
19 words
A method according to claim 21, wherein the diameter of the microspheres is a minimum of about 10 μm.
Claim 53
Dependent Claim
19 words
A method according to claim 22, wherein the diameter of the microspheres is a minimum of about 10 μm.
Claim 54
Dependent Claim
19 words
A method according to claim 23, wherein the diameter of the microspheres is a minimum of about 10 μm.
Claim 55
Dependent Claim
19 words
A method according to claim 28, wherein the diameter of the microspheres is a minimum of about 10 μm.
References Cited (Backward Citations)
US4266032A
Cited
USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
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| 3.5 Years (Year 4) | 1995-05-19 | 1995-11-18 | 1996-05-19 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1999-05-19 | 1999-11-18 | 2000-05-19 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 2003-05-19 | 2003-11-18 | 2004-05-19 | $7700.00 | Window Closed / Lapsed |