Wednesday, January 4, 2012

Isolation of murine alveolar macrophages by lung lavage

Phagocytosis by alveolar macrophages (AM) is the primary defense mechanism against micro-organisms in the lung. Alveolar macrophages can also be a potent suppressors of T lymphocyte responses, which is important for the limitation of tissue damage. Alveolar macrophages reside in an environment that is quite different than most macrophages due to the aerobic atmosphere. 
In this protocol we describe the method for for isolation of murine alveolar macrophages by lung lavage . 

Method:
1) Sacrifice the mice and bleed them by aortic section.
2) Excise the lungs from the thoracic cavity and place the lungs on a sterile Petri dish. Work under a sterile hood.
3) Cannulate the trachea with a 18-gauge needle and lavage the lungs 3 times with sterile saline. Use 35 ml/kg of body weight. For a 25 gram mouse use 0.875 ml of sterile saline warmed to 37 C for each lavage .
4) Collect the cell suspension in a conical tube and centrifuge at 450 g for 10-15 minutes.
5) Resuspend in RPMI 1640 containing 10% fetal calf serum (FCS). 
RPMI 1640 can be purchased from Invitrogen:

For more information about macrophage depletion visit www.clodrosome.com

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