Harvest and Culture of Mouse Peritoneal Macrophages
小鼠腹水中巨噬细胞的采集和培养
发布: 2013年11月20日第3卷第22期 DOI: 10.21769/BioProtoc.976 浏览次数: 39436
评审: Fanglian He
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参见作者原研究论文
The authors used this protocol in:
May 2013
Abstract
Peritoneal macrophages are used as primary macrophages in lots of studies, mainly because they are easy to obtain. Injection of thioglycollate broth i.p. induces inflammatory responses and elicits large numbers of macrophages. This protocol can be used for harvesting resident or thioglycollate-elicited peritoneal cells. Peritoneal macrophages are non-adherent in situ and when they are cultured in dishes, they become adherent so that macrophages may be separated from other types of cells in peritoneal cavity.
Materials and Reagents
- C57BL/6 mouse
- 70% alcohol or isopropanol
- PBS (Life Technologies, catalog number: 10010023 )
- EDTA (Life Technologies, catalog number: 15575-020 )
- RPMI 1640 (Cellgro®, catalog number: 10-041-CV )
- 10% heat-inactivated FBS (endotoxin < 0.06 EU/ml) (Hyclone, catalog number: SH30071.03HI )
- Penicillin-Streptomycin-Glutamine (Life Technologies, catalog number: 10378016 )
- Nonessential amino acids (Life Technologies, catalog number: 11140050 )
- Sodium pyruvate (Sigma-Aldrich, catalog number: S8636 )
- Hepes (suitable for cell culture) (Sigma-Aldrich, catalog number: H0887 )
- 2-mercaptoethanol (EMD Millipore, catalog number: ES-007-E )
- Thioglycollate medium (BD Biosciences, catalog number: 211716 )
- Pyrogen-free water (Hyclone, catalog number: SH30529.03 )
- cRPMI medium (see Recipes)
Equipment
- 5 ml syringe
- 12-well plate
- 15 ml sterile Falcon tubes
- Needles (20 G and 25 G)
- Centrifuge
- 37 °C, 5% CO2 cell culture incubator
- 300 ml flask with an aluminum foil lid
- Laboratory oven
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:
- Lu, M. and Varley, A. W. (2013). Harvest and Culture of Mouse Peritoneal Macrophages. Bio-protocol 3(22): e976. DOI: 10.21769/BioProtoc.976.
- Lu, M., Varley, A. W. and Munford, R. S. (2013). Persistently active microbial molecules prolong innate immune tolerance in vivo. PLoS Pathog 9(5): e1003339.
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分类
细胞生物学 > 细胞分离和培养 > 细胞分离
免疫学 > 免疫细胞分离 > 巨噬细胞
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