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Preparation of BMMCs and BMDMs

SW Steven S. Welc
IF Ivan Flores
MW Michelle Wehling-Henricks
JR Julian Ramos
YW Ying Wang
CB Carmen Bertoni
JT James G. Tidball
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BMMCs were separated from whole BMC preparations flushed from WT or CD11b/LIF femurs and tibiae and separated using a histopaque-1077 gradient (Sigma). The freshly-isolated BMMCs were then used for RNA isolation and analysis. For preparation of BMDMs, BMCs were aseptically flushed from WT or CD11b/LIF femurs and tibiae and differentiated in vitro to BMDMs24. BMDMs were stimulated for 24 h with activation media consisting of Dulbecco’s Modified Eagle Medium (DMEM) with 0.25% heat-inactivated fetal bovine serum (FBS; Omega), 100 U/ml penicillin, 100 µg/ml streptomycin (1% P/S), and 10 ng/ml macrophage colony stimulating factor (MCSF; R&D).

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