Primary Culture of SVZ-derived Progenitors Grown as Neurospheres
室管膜下区(SVZ)源祖细胞生长为神经球的初代培养
发布: 2013年08月20日第3卷第16期 DOI: 10.21769/BioProtoc.868 浏览次数: 13283
评审: Xuecai Ge
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参见作者原研究论文
The authors used this protocol in:
Feb 2013
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Abstract
SVZ-derived progenitors grown as neurospheres is a well-known model to study neural stem cell and progenitor functions such as proliferation, differentiation/self-renewal, and migration (Durbec and Rougon, 2001). This protocol is for preparing a culture of SVZ-derived progenitors from 8 early postnatal mouse brains (P0 to P3). One week after cell plating, we can observe round floating neurospheres, each resulting from the clonal expansion of a single EGF/FGF responsive neural progenitor.
Keywords: Nervous system (神经系统)
Adult brain (成人的大脑)
Stem cells (干细胞)
Subventricular zone (室管膜下区)
Primary culture (原代培养)
Materials and Reagents
- New born mice
- Phosphate Buffered Saline (PBS) (Life Technologies, catalog number: 14040-091 )
- Hank's Balanced Salt Solution (HBSS) (Life Technologies, catalog number: 14170-088 )
- Dulbecco's Modified Eagle Medium (DMEM) (Life Technologies, catalog number: 61965-026 )
- Ham’s F-12 nutrient mix (F12) (Life Technologies, catalog number: 31765-027 )
- Trypsin (Sigma-Aldrich, catalog number: T5266 )
- Insulin (Sigma-Aldrich, catalog number: I1882 )
- Holo-transferrin (Sigma-Aldrich, catalog number: T0665 )
- Putrescine (Sigma-Aldrich, catalog number: P5780 )
- Progesterone (Sigma-Aldrich, catalog number: P8783 )
- Sodium selenite (Selenium) (Sigma-Aldrich, catalog number: S5261 )
- Penicillin-streptomycin (Life Technologies, catalog number: 15140-130 )
- Fetal Bovine Serum (FBS) (Life Technologies, catalog number: 10106-169 )
- B27 (Life Technologies, catalog number: 17504-044 )
- rhFGFbasic (Peprotech, catalog number: 167 100-18B-B )
- rhEGF (Peprotech, catalog number: 167 AF-100-15A )
- Neurospheres defined medium (see Recipes)
- Trypsin (see Recipes)
Equipment
- Sterilin Universal Container (Thermo Fisher Scientific, catalog number: 128A/P )
- Standard TC BD Falcon 60 mm cell culture dish (BD Biosciences, Falcon®, catalog number: 353002 )
- Vibratom (Microm, model: HM450 )
- Stereoscopic Microscope
- Cell culture Hood and Incubator
- Scissors, fine forceps, spatula, micro knives
- Ice bucket
- Fire-polished glass Pasteur pipettes
- Water bath
Procedure
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文章信息
版权信息
© 2013 The Authors; exclusive licensee Bio-protocol LLC.
如何引用
Readers should cite both the Bio-protocol article and the original research article where this protocol was used:
- Vernerey, J., Magalon, K. and Durbec, P. (2013). Primary Culture of SVZ-derived Progenitors Grown as Neurospheres. Bio-protocol 3(16): e868. DOI: 10.21769/BioProtoc.868.
- Vernerey, J., Macchi, M., Magalon, K., Cayre, M. and Durbec, P. (2013). Ciliary neurotrophic factor controls progenitor migration during remyelination in the adult rodent brain. J Neurosci 33(7): 3240-3250.
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分类
神经科学 > 发育 > 神经元
干细胞 > 成体干细胞 > 神经干细胞
细胞生物学 > 细胞分离和培养 > 细胞分化
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