Stem Cell Aggregate Suspension Compositions and Methods of Differentiation Thereof
First Claim
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1. A method of generating a hES-derived cell aggregate in suspension from a hES single cell suspension, comprising:
- (a) culturing a hES cell in ani adherent growth culture condition which allows for expansion in an undifferentiated state;
(b) disassociating the adherent hES cell culture in (a) into a hES single cell suspension culture; and
(d) contacting the hES single cell suspension culture with a differentiating culture condition which allows for formation of hES-derived cell aggregates in suspension when agitated until such a period of time when the single cell suspension culture forms a hES-derived cell aggregate in suspension,thereby generating a hES-derived cell aggregate in suspension.
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Abstract
The present invention relates to methods for production of undifferentiated or differentiated embryonic stem cell aggregate suspension cultures from undifferentiated or differentiated embryonic stem cell single cell suspensions and methods of differentiation thereof.
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Citations
24 Claims
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1. A method of generating a hES-derived cell aggregate in suspension from a hES single cell suspension, comprising:
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(a) culturing a hES cell in ani adherent growth culture condition which allows for expansion in an undifferentiated state; (b) disassociating the adherent hES cell culture in (a) into a hES single cell suspension culture; and (d) contacting the hES single cell suspension culture with a differentiating culture condition which allows for formation of hES-derived cell aggregates in suspension when agitated until such a period of time when the single cell suspension culture forms a hES-derived cell aggregate in suspension, thereby generating a hES-derived cell aggregate in suspension. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18)
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19. A method of generating a hES-derived cell aggregate in suspension from a hES-derived single cell suspension, comprising:
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(a) culturing a hES cell in an adherent growth culture condition which allows for expansion in an undifferentiated state; (b) contacting the undifferentiated hES cell with a first differentiating culturing condition suitable for differentiating the hES cell and resulting in an adherent hES-derived cell; (c) disassociating the adherent hES-derived cell into a single cell suspension culture; and (d) contacting the single cell suspension culture with a second differentiating culture condition which allows for formation of hES-derived cell aggregates in suspension when agitated until such a period of time when the single cell suspension culture forms a hES-derived cell aggregate in suspension, thereby generating a hES-derived cell aggregate in suspension. - View Dependent Claims (20)
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21. A method for enriching pancreatic progenitor cells comprising using low concentrations of growth factors selected from the group consisting of FGF10, EGF, KGF, noggin and retinoic acid.
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22. A method for enriching endocrine progenitor cells comprising using high concentrations of growth factors selected from the group consisting of FGF10, EGF, KGF, noggin and retinoic acid.
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23. A method for enriching pancreatic progenitor cells from a hES cell culture comprising increasing the hES cell density culture from which is derived the pancreatic progenitor.
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24. A method for enriching endocrine progenitor cells from a hES cell culture comprising decreasing the hES cell density culture from which is derived the pancreatic progenitor.
Specification