Dulbecco s modified eagle s media

The mouse macrophage cell line RAW 264.7 was obtained from the American Type Culture Collection. The cells were cultured in Dulbecco’s modified Eagle’s media (WELGENE, Daegu, Korea), which contained 10% fetal bovine serum (HyClone, Logan, UT, USA) and 1% penicillin/streptomycin (Gibco, Grand Island, NY, USA). The cells were cultured at 37 °C in an incubator with a humidified atmosphere of 5% CO₂.

HEK293T cells were purchased from American Type Culture Collection (Manassas,VA), cultured in Dulbecco’s modified Eagle’s media (Welgene, Daegu, Korea) supplemented with 10% fetal bovine serum (Thermo Fisher Scientific Korea, Seoul, Korea), and transfected with plasmid using Lipofectamine 2000 (Thermo Fisher Scientific Korea) according to the manufacturer’s instructions. Subsequently, the cells were lysed with M-PER mammalian protein extraction reagent (Thermo Fisher Scientific Korea) at 48 h post-transfection, and processed for Western blotting as described previously [27 (link)]. The antibodies used were anti-FLAG antibody (1:2000, Sigma-Aldrich, catalog number F1804), anti-glyceraldehyde-3-phosphate dehydrogenase (anti-GAPDH) antibody (1:2000, Trevigen, Gaithersburg, MD, 2275-PC-100), HRP-conjugated goat anti-mouse antibody (1:4000, Santa Cruz Biotechnology, Dallas, TX, sc-2005), and HRP-conjugated goat anti-rabbit antibody (1:4000, Santa Cruz Biotechnology, sc-2004). Band intensity on the Western blots was analyzed using ImageJ.

The HeLa cell line was obtained from the Korean Cell Line Bank (KCLB). The HeLa cells were cultured in Dulbecco's modified Eagle's media (Welgene, Daegu, Korea) supplemented with 10% heat-activated fetal bovine serum (Biowest, Nuaillé, France) and 1% penicillin or streptomycin mixtures (GIBCO, ThermoFisher, MA, USA). The HeLa cells were incubated in a humidified atmosphere at a CO₂ concentration of 5% and a temperature of 37°C.