Bone Marrow-Derived Macrophages isolation and differentiation

Download PDF

Scope of the method

The Method relates to
  • Human health
The Method is situated in
  • Basic Research
Type of method
  • In vitro - Ex vivo
This method makes use of
  • Animal derived cells / tissues / organs
Species from which cells/tissues/organs are derived
Mus musculus - C57BL/6J
Type of cells/tissues/organs
Bone marrow derived macrophages

Description

Method keywords
  • BMDMs
  • macrophage polarization
  • macrophages
  • isolation
  • Culturing
Scientific area keywords
  • Immunometabolism
  • Immunology
  • neurodegeneration
  • neuroinflammation
Method description

Isolation Femoral and tibial bones are isolated from 10-12 weeks old C57BL/6J mice. After cutting the bone's ends, the bone marrow is flushed out using a 23G syringe with PBS into a 50ml falcon tube. After centrifugation cells are re-suspended in FCS (0.5ml/mouse leg). Differentiation 0.5ml of the cells suspension is added to 20ml BMM medium (15% LCM, 10% FCS and 0.5% P/S in RPMI1640) in a 20cm petri dish. After 5 days of culture all medium is replaced by 20ml fresh BMM medium. On day 7 cells can be plate out for experiments.

Lab equipment
  • - Biosafety cabinet flow hood ;
  • - 5% CO2 Incubator ;
  • - Centrifuge ;
  • - Dissection material.
Method status
  • History of use
  • Internally validated
  • Published in peer reviewed journal

Pros, cons & Future potential

Advantages
  • - High amount of cells per isolation ;  
  • - Possibility of freezing the cells for future use after isolation ; 
  • - Reproducibility.
Modifications

If the cells are confluent at day 5, instead of discard the BMM medium you can place it into a new petridish to increase the amount of cells.

References, associated documents and other information

References

Bogie JF, Mailleux J, Wouters E, Jorissen W, Grajchen E, Vanmol J, Wouters K, Hellings N, Van Horsen J, Vanmierlo T, et al. 2017b. Scavenger receptor collectin placenta 1 is a novel receptor involved in the uptake of myelin by phagocytes. Sci Rep 7: 44794.

Contact person

Jerome Hendriks

Organisations

University of Hasselt (UHasselt)
Biomedisch Onderzoeksinstituut
Belgium
Flemish Region