Culturing Mouse and Human Macrophages

Mouse iBMDMs were gifts from Dr. Jonathan Kagan. Cells were cultured in Dulbecco’s Modified Eagle’s medium (DMEM, Corning 15013CV) supplemented with 10% fetal bovine serum (FBS, Corning 35–016-CV), 2 mM L-glutamine (Corning, MT25005CI), and 0.1% penicillin/streptomycin (P/S, Corning, MT3000CI) at 37°C in humidified incubators with 5% CO₂. Cells were passaged every 2–3 days when they reached 60%–80% confluency. Human embryonic kidney 293T (HEK293T, ATCC, CRL3216) cells were cultured in DMEM supplemented with 10% FBS, 2 mM L-glutamine, and 0.1% penicillin/streptomycin. Human THP1 macrophages (ATCC, TIB202) were cultured in RPMI 1640 (ATCC modification, Gibco A1049101) supplemented with 10% FBS and differentiated with 100 nM phorbol 12-myristate 13-acetate (PMA, Sigma-Aldrich, P8139) for 24 hours. Cell lysate of primary mouse bone marrow-derived macrophages was a gift from the laboratory of Dr. Yuan He at Wayne State University (66 (link)).