Protein Flotation Assay to Isolate Lipids Rafts from Soft Tissue or Cells
蛋白质浮选分离试验从软组织或细胞中分离脂筏
发布: 2013年08月20日第3卷第16期 DOI: 10.21769/BioProtoc.854 浏览次数: 11902
评审: Xuecai Ge
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参见作者原研究论文
The authors used this protocol in:
Jan 2013
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Abstract
The arrangement in eukaryotic cell membranes of liquid-ordered states surrounded by liquid-disordered phases allows for the existence of organized membrane microdomains called rafts. Rafts play a crucial role in signal-transduction events, in lipid transport and in various internalization processes, and for all these reasons need to be purified for further characterization.
Materials and Reagents
- Optiprep® (Optiprep® is a 60% solution of iodixanol in water, density = 1.32 g/ml) (Sigma-Aldrich, catalog number: 079K1726 )
- Sodium chloride (NaCl)
- Tris hydrochloride (Tris-HCl)
- Ethylenediaminetetraacetic acid (EDTA) (Sigma-Aldrich, catalog number: ED255 )
- Triton X-100 (Sigma-Aldrich, catalog number: T-8532 )
- Methanol/chloroform
- Lysis buffer (see Recipes)
- Gradient buffer (see Recipes)
Equipment
- Pellet pestles (Sigma-Aldrich, catalog number: Z359963-1EA )
- Sorvall® WX90 ultracentrifuge (Thermo Fisher Scientific, catalog number: 46901 )
- Sorvall® TH-641 swinging bucket rotor (Thermo Fisher Scientific, catalog number: 54295 )
- Ultracentrifuge tubes: Beckman Coulter® open-top Ultra-ClearTM tubes (dimension: 14 x 89 mm, nominal volume: 12 ml) (Beckman Coulter®, catalog number: 344059 )
- 1 ml pipette
- Western Blotting apparatus
Procedure
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:
- Lemaire-Vieille, C., Gagnon, J. and Cesbron, J. (2013). Protein Flotation Assay to Isolate Lipids Rafts from Soft Tissue or Cells. Bio-protocol 3(16): e854. DOI: 10.21769/BioProtoc.854.
- Lemaire-Vieille, C., Bailly, Y., Erlich, P., Loeuillet, C., Brocard, J., Haeberle, A. M., Bombarde, G., Rak, C., Demais, V., Dumestre-Perard, C., Gagnon, J. and Cesbron, J. Y. (2013). Ataxia with cerebellar lesions in mice expressing chimeric PrP-Dpl protein. J Neurosci 33(4): 1391-1399.
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分类
生物化学 > 蛋白质 > 分离和纯化
生物化学 > 脂质 > 脂筏
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