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查看更多产品信息 Rat Fetal Neural Stem Cells - FAQs (N7744100)
26 个常见问题解答
这些细胞是从E14孕期的Sprague Dawley大鼠胚胎皮层中分离得到的。它们能够以未分化的性状在培养条件下扩增三代。
以下生长因子可用于NSC的扩增：重组EGF（货号PHG0314），重组bFGF（货号PHG0024)，和重组VEGF（货号PHC9394）。此外，包括BDNF在内的多几种神经营养因子例如BDNF（货号10908010），CNTF（货号PHC7015），和GDNF（货号PHC7044）也可适用于相关研究。
人NSC可培养于Gibco StemPro NSC SFM（货号A1050901）和Gibco Geltrex 基质/或Gibco CELLstart 底物基质预先包被的培养皿中。另外，如果用户的研究目的是获得神经元，也可在已添加了Gibco B-27添加剂（不含维生素A）的Neurobasal培养基（不含维生素A） + 预先包被的培养皿中培养NSC。
当按照克隆密度进行培养时，通常可基于用神经球的形成能力来鉴定NSC（Nat Methods 2:333 (2005)）。也可通过（1）Sox1，Sox2和Nestin的RT-PCR检测或（2）nestin，Pax6，Sox2和Ki67的免疫组化染色来鉴定NSC。
神经干细胞（NSC）是神经系统中能够自我更新的多专能细胞，拥有分化为神经元，少突胶质细胞和星形胶质细胞的潜能。神经干细胞可由胚胎干细胞诱导分化而来，或从皮层，脑室下区（SVZ）和脑室区等多个不同脑区分离获得，或从骨髓来源的间充质干细胞（MSC）生成 (J Cell Biochem 114:764 (2013))。NSC是研究神经发生和神经递质与受体功能的宝贵工具。来自在各类动物疾病模型中的NSC被广泛应用于包括例如帕金森氏病与亨廷顿氏病舞蹈症在内的等多种不同类型的CNS疾病的研究中（J Cell Biochem 114:764 (2013)）。
If you are growing cells on cover slips to do immunocytochemistry, we recommend using Permanox chamber slides:
- Permanox chamber slides, 4-well, pk/16 (VWR, Cat. No. 62407-330)
- Permanox chamber slides, 8-well, pk/16 (VWR, Cat. No. 62407-335)
If you are growing cells on cover slips for other purposes and require coating, use this procedure to double coat the cover slips:
- Incubate cover slips overnight at room temperature in poly-ornithine solution (final concentration = 100 mg/mL in cell culture water)
- Rinse twice with sterile water
- Incubate cover slips overnight at 4 degrees C in laminin solution (final concentration = 15 mg/mL in cell culture water)
Based on the expression of specific phenotypic markers, here are the expected percentages of the different cell populations:
Dcx marker (neuronal): 10% positive with spontaneous differentiation; 40% positive with directed differentiation
GFAP marker (astrocyte): 30% positive with spontaneous differentiation; 40% positive with directed differentiation
O4 marker (oligodendrocyte): 2% positive with spontaneous differentiation; 32% positive with directed differentiation
Cortical NSC's contain a greater mixture of cells and have different neurotransmitters. They have a wider set of applications. Fewer NSCs can be obtained from hippocampal tissue, but the cells are more pure. They are used by a smaller hippocampal-focused audience
Gibco Rat Fetal Neural Stem Cells are derived from a mixed pool of embryos; therefore, cells will be derived from rats of both sexes.
Any format of culture vessel (e.g., flask, 6-well plates, etc.) can be used. Seed cells at a density of 5 x 10E4 cells/cm2. Depending on the surface area of your culture vessel, calculate how many cells you will need to have on hand. Example: For one 60 mm dish, you will need 5 x 10E4 cells/cm2 x 20 cm2 = 1 x 10E6 cells total.
You can find protocols for directed and spontaneous differentiation of Rat Neural Stem Cells in the Neurobiology Protocol Handbook on our website: https://assets.thermofisher.com/TFS-Assets/BID/Handbooks/gibco-neurobiology-protocol-handbook.pdf
Neural stem cells (NSC) is a valuable source not only for neuroscience but also for clinical use to treat neurodegenerative disease or neurological disorder. These cells can be used for a wide range of differentiation studies.
Adult derived NSCs tends to be regionally restricted and have limited plasticity compared to fetal derived NSCs.
To maintain the cells in NSC status, we recommend using NSC-SFM. We have validated and optimized the culture conditions in NSC-SFM to ensure superior growth and performance of these cells.
Please ensure that the buffer used for CELLstart has Calcium and Magnesium. We recommend using Dulbecco's Phosphate Buffered Saline (D-PBS), containing calcium and magnesium, but no phenol red (Cat. No. 14040133). If the right buffer is used, try increasing the concentration of CELLstart up to 1:50 dilution rate. If CELLstart with our recommended buffer does not work, you can try using fibronectin or poly-L-ornithine-coated plates. If you use poly-L-orinithine, please perform an overnight incubation.
The doubling time for the cells is around 20-30 hrs. This tends to increase with passage number.
Upon thawing, the vial should have more than 50% viability which will provide more than 2 million live cells.
Cell purity was measured by testing for the expression of the NSC phenotype marker, Nestin and the lack of expression of differentiated phenotype markers such as GFAP, Dcx, or GalC. More than 75% of the cells were positive for Nestin and expression of the differentiated markers was less than 10%.
We evaluated Gibco Rat Fetal Neural Stem Cells for two criteria:
- To express the NSC phenotype marker, Nestin and not express differentiated phenotype markers such as GFAP, Dcx, or GalC
- To demonstrate their differentiation potential toward all three downstream lineages such as neurons, astrocytes, and oligodendrocytes
Rat NSC have been tested for proliferation and differentiation potential up to passage 3 after thawing and we have seen around 300 million cells from 1 vial after the 3rd passage. We have not expanded the cells beyond the third passage, but we encourage you to try expanding further.
Cells were isolated from the fetus without passage and frozen down at passage 0.
These cells were isolated from the cortex of Sprague-Dawley rats at day E14 of gestation. They can be expanded in culture up to 3 passages without differentiation.
Find additional tips, troubleshooting help, and resources within our Cell Culture Support Center.
For NSC expansion, the following growth factors are used: recombinant EGF (Cat. No. PHG0314), recombinant FGF-basic (Cat. No. PHG0024), and recombinant VEGF (Cat. No. PHC9394). In addition, several neurotrophins such as BDNF Cat. No. 10908010), NT-3, CNTF (Cat. No. PHC7015), and GDNF (Cat. No. PHC7044)are also used in the related studies.
Information pertaining to whether a specific product has been tested against the WHO Reference Standard can typically be located on the product page or Certificate of Analysis (COA).
Find additional tips, troubleshooting help, and resources within our Cell Culture Support Center.
Human NSCs can grow in Gibco StemPro NSC SFM (Cat. No. A1050901) on dishes pre-coated with Gibco Geltrex Matrix or Gibco CELLstart substrate. Alternatively, if the goal is to obtain neurons, NSCs can also be grown on Neurobasal medium supplemented with Gibco B-27 supplements without vitamin A on a pre-coated dish.
NSCs are generally characterized by their ability to form neurospheres when plated at cloning density (Nat Methods 2:333 (2005)). NSCs can also be characterized by (1) RT-PCR of Sox1, Sox2, and Nestin or (2) immunohistochemical staining for nestin, Pax6, Sox2, and Ki67.
Neural stem cells (NSCs) are self-renewing multipotent cells of the nervous system capable of differentiating into neurons, oligodendrocytes, and astrocytes. NSC can be generated by induced differentiation from embryonic stem (ES) cells, or isolated from various regions of the brain including the cortex, the subventricular zone (SVZ), and the ventricular zone, or generated from bone marrow-derived mesenchymal stem cells (MSCs) (J Cell Biochem 114:764 (2013)). NSCs are valuable tools for the study of neurogenesis and neurotransmitter and receptor function. NSCs were used in the investigation of different CNS disorders such as PD and Huntington's disease in various animal models (J Cell Biochem 114:764 (2013)).