Medium for growing human embryonic stem cells
First Claim
1. A method for proliferating hES cells so that they do not differentiate, comprising:
- obtaining an hES cell culture medium, and then culturing hES cells in the medium in the presence of an extracellular matrix and/or feeder cells so that the hES cells proliferate without differentiation;
wherein said hES cell culture medium comprises;
a) an isotonic buffer;
b) a protein nutrient, comprising serum, serum replacement, albumin, or essential and non-essential amino acids;
c) lipids, fatty acids, or cholesterol, either as artificial additives or as the HDL or LDL extract of serum;
d) fibroblast growth factor at a concentration of at least 40 ng/mL.
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Abstract
This disclosure provides an improved system for culturing human pluripotent stem cells. Traditionally, pluripotent stem cells are cultured on a layer of feeder cells (such as mouse embryonic fibroblasts) to prevent them from differentiating. In the system described here, the role of feeder cells is replaced by components added to the culture environment that support rapid proliferation without differentiation. Effective features are a suitable support structure for the cells, and an effective medium that can be added fresh to the culture without being preconditioned by another cell type. Culturing human embryonic stem cells in fresh medium according to this invention causes the cells to expand surprisingly rapidly, while retaining the ability to differentiate into cells representing all three embryonic germ layers. This new culture system allows for bulk proliferation of pPS cells for commercial production of important products for use in drug screening and human therapy.
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Citations
12 Claims
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1. A method for proliferating hES cells so that they do not differentiate, comprising:
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obtaining an hES cell culture medium, and then culturing hES cells in the medium in the presence of an extracellular matrix and/or feeder cells so that the hES cells proliferate without differentiation;
wherein said hES cell culture medium comprises;
a) an isotonic buffer;
b) a protein nutrient, comprising serum, serum replacement, albumin, or essential and non-essential amino acids;
c) lipids, fatty acids, or cholesterol, either as artificial additives or as the HDL or LDL extract of serum;
d) fibroblast growth factor at a concentration of at least 40 ng/mL. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10)
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11. A method for proliferating hES cells so that they do not differentiate, comprising:
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obtaining an hES cell culture medium and an extracellular matrix, and then culturing hES cells in the medium on the extracellular matrix so that the hES cells proliferate without differentiation;
wherein said hES cell culture medium comprises;
a) an isotonic buffer;
b) a protein nutrient, comprising serum, serum replacement, albumin, or essential and non-essential amino acids;
c) lipids, fatty acids, or cholesterol, either as artificial additives or as the HDL or LDL extract of serum;
d) fibroblast growth factor at a concentration of at least 40 ng/mL. - View Dependent Claims (12)
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Specification