*2.5. Isolation and Generation of Bone Marrow-Derived Macrophages (BMMΦs) and Dendritic Cells (BMDCs)*

BMMΦs [24] and BMDCs [25] were generated from isolated bone marrow as previously described. Briefly, 5–12-week C57BL/6J female mice were euthanized and the femurs and tibias isolated and flushed with BMMΦ media [RPMI 1640 supplemented with L-glutamine (Life Technologies, Carlsbad, CA), penicillin (100 units/mL), streptomycin (100 µg/mL), 10% heat-inactivated fetal bovine serum (FBS) (VWR, Radnor, PA, USA), and 20% L929 (ATCC) cell-conditioned media] or BMDC media [RPMI 1640 supplemented with L-glutamine, penicillin (100 units/mL), streptomycin (100 µg/mL), 10% FBS, 50 mM β-mercaptoethanol (Sigma-Aldrich) and 20 ng/mL GM-CSF (Peprotech, Rocky Hill, NJ, USA)] using a 1 mL syringe and a 25-gauge needle. Once isolated, the cells were pipetted and filtered through a 40 µm cell strainer then plated in uncoated 10 cm non-tissue culture treated petri dishes. The cells were incubated at 37 ◦C at 5% CO<sup>2</sup> and the media was

replaced on days 0, 3, 6, and 8. BMMΦs and BMDCs were used for experiments between days 8–10.
