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Serum free cultivation of primate embryonic stem cells

a technology embryonic stem cells, which is applied in the field of serum free cultivation of primate embryonic stem cells, to achieve the effects of avoiding variability in results due to differences in animal serum batches, avoiding use of animal serum, and increasing the efficiency of cloning

Inactive Publication Date: 2006-02-23
THOMSON JAMES A
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

"The invention provides methods for culturing primate embryonic stem cells in a way that is less variable and allows for more efficient cloning. The methods involve culturing the stem cells in a culture that is free of mammalian fetal serum and in the presence of fibroblast growth factor, which is supplied from a source other than just a fibroblast feeder layer. The culturing can be done with a fibroblast feeder layer or with a growth factor that activates a fibroblast growth factor signaling receptor. The methods also involve using a culture system that has a fibroblast feeder layer and human basic fibroblast growth factor supplied by other than just the fibroblast feeder layer. The invention also provides cell lines derived using these methods. Overall, the invention provides more stable and consistent conditions for culturing primate embryonic stem cells."

Problems solved by technology

However, when one or more well defined purified component(s) of such serum is added, they do not.

Method used

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Embodiment Construction

[0023] In the following experiments I used the methods and culture systems of the invention to culture human ES cell lines. Two clonally derived human ES cell lines proliferated for over eight months after clonal derivation and maintained the ability to differentiate to advanced derivatives of all three embryonic germ layers.

[0024] Techniques for the initial derivation, culture, and characterization of the human ES cell line H9 were described in J. Thomson et al., 282 Science 1145-1147 (1998). In my experiments herein human ES cells were then plated on irradiated (35 gray gamma irradiation) mouse embryonic fibroblast. Culture medium for the present work consisted of 80% “KnockOut” Dulbeco's modified Eagle's medium (DMEM) (Gibco BRL, Rockville, Md.), 1 mM L-Glutamine, 0.1 mM β-mercaptoethanol, and 1% nonessential amino acids stock (Gibco BRL, Rockville, Md.), supplemented with either 20% fetal bovine serum (HyClone, Logan, Utah) or 20% KnockOut SR, a serum-free replacement originall...

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Abstract

Disclosed herein are methods for culturing primate embryonic stem cells. These cells are cultured on a prolonged and stable basis in the presence of exogenously supplied fibroblast growth factor and in the absence of animal serum. Preferably there is also a fibroblast feeder layer. Also disclosed is a culture media containing fibroblast feeder layer and the fibroblast growth factor.

Description

CROSS REFERENCES TO RELATED APPLICATIONS [0001] Not applicable. STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH BACKGROUND OF THE INVENTION [0002] The present invention relates to methods for culturing primate embryonic stem cell cultures and culture media useful therewith. [0003] Primate (e.g. monkey and human) pluripotent embryonic stem cells have been derived from preimplantation embryos. See U.S. Pat. No. 5,843,780 and J. Thomson et al., 282 Science 1145-1147 (1998). The disclosure of these publications and of all other publications referred to herein are incorporated by reference as if fully set forth herein. Notwithstanding prolonged culture, these cells stably maintain a developmental potential to form advanced derivatives of all three embryonic germ layers. [0004] Primate (particularly human) ES cell lines have widespread utility in connection with human developmental biology, drug discovery, drug testing, and transplantation medicine. For example, current knowledge of the ...

Claims

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Application Information

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Patent Type & Authority Applications(United States)
IPC IPC(8): A01K67/00C12N5/06C12N5/08A61K9/22C12N5/07A61K31/495C12NC12N5/00C12N5/02C12N5/0735C12R1/91
CPCC12N5/0606C12N2501/115C12N2500/99C12N2500/90C12N5/06
Inventor THOMSON, JAMES A.
Owner THOMSON JAMES A