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US 2009/0263901 A1Published application
Stable neural stem cell lines
AssigneeNeuralStem Biopharmaceuticals, Ltd.
InventorsRenji Yang, Karl K. Johe
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Abstract
A systematic and efficient method for establishing stable neural stem cell lines and neuronal progenitor lines is described. The resulting cell lines provide robust, simple, and reproducible cultures of human and other mammalian neurons in commercially useful mass quantities while maintaining normal karyotypes and normal neuronal phenotypes.
Claims22
A method for producing stable cell lines of mammalian neural precursor cells in vitro, comprising the steps of:
- a) preparing a culture of neural precursor cells in a serum-free medium;
- b) culturing the neural precursor cells in the presence of a first mitogen, wherein said first mitogen is selected from the group consisting of aFGF, bFGF, EGF, TGFα and combinations thereof;
- c) contacting the cells with an agent capable of being taken up by the cells and capable of expressing a c-myc gene;
- and
- d) further culturing the cells in a medium containing the first mitogen and a second mitogen, wherein said second mitogen is selected from the group consisting of aFGF, bFGF, EGF, TGFα serum and combinations thereof, with the proviso that the second mitogen is other than the first mitogen.
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A method for producing stable clonal cell lines of mammalian neural precursor cells in vitro, comprising the steps of:
- a) preparing a culture of neural precursor cells in a serum-free medium;
- b) culturing the neural precursor cells in the presence of a first mitogen, wherein said first mitogen is selected from the group consisting of aFGF, bFGF, EGF, TGFα and combinations thereof;
- c) contacting the cells with an agent capable of being taken up by the cells and capable of expressing a c-myc gene and a selectable marker;
- d) further culturing the cells in a medium containing the first mitogen and a second mitogen, wherein said second mitogen is selected from the group consisting of aFGF, bFGF, EGF, TGFα, serum and combinations thereof, with the proviso that the second mitogen is other than the first mitogen;
- and
- e) collecting c-myc treated cells and co-culturing them with feeder cells free of the selectable marker and capable of supporting survival of the c-myc treated cells in a medium containing the first mitogen and the second mitogen, with the proviso that the second mitogen is other than the first mitogen.
3 more elements
Dependencies are inferred from the claim text.