Non coated petri dishes

BMCs isolated from C57BL/6J mice were used for bone marrow macrophage and osteoclast differentiation. Brie y, BMCs were ushed from tibiae and femorae. After the lysis of red blood cells, the BMCs were plated in a 10 cm tissue culture dish with α-MEM complete medium (basal α-MEM medium supplemented with 10% FBS, 100 U/ml penicillin G, and 100 µg/ml streptomycin and 2 mM glutamine) and 20 ng/ml mM-CSF (macrophage/monocyte colony stimulating factor). After an overnight incubation in a humidi ed atmosphere of 5% CO 2 at 37 °C, the non-adherent BMCs were collected for further experimentation.
2.5 RAW264.7 cell culture RAW264.7 cells were purchased from American Type Culture Collection (ATCC, Manassas, VA). The RAW264.7 cells were maintained in high glucose D-MEM complete medium (basal D-MEM supplemented with 10% fetal bovine serum, 100 U/ml penicillin G, and 100 µg/ml streptomycin and 2 mM glutamine), and in a humidi ed atmosphere of 5% CO 2 at 37 °C. Cells were maintained in suspension culture in noncoated petri dishes (Greiner Bio-one, Monroe, NC), which markedly reduces drift toward a differentiated state. For experiments cells were plated on standard tissue culture dishes.