Recombinant mouse interlukin 4

BMDCs were isolated and differentiated from monocytes as previously described, with minor modifications [13 (link)]. Briefly, femurs and tibiae from 6-week-old C57BL/6 mice (Raon Bio, Yongin, Kyonggi-do, Korea) were isolated and flushed with complete RPMI media to collect bone marrow. Red blood cells were lysed to obtain monocyte pellets, which were suspended and cultured in Iscove’s modified Dulbecco’s medium supplemented with 10% fetal bovine serum (FBS; GenDEPOT, Barker, TX, USA), 100 units/mL penicillin, 100 mg/mL streptomycin (Gibco, Carlsbad, CA, USA), 20 ng/mL recombinant mouse granulocyte-macrophage colony-stimulating factor, 20 ng/mL recombinant mouse interlukin-4 (GenScript, Piscataway, NJ, USA), and 50 μM β-mercaptoethanol (Sigma-Aldrich). The medium was changed every 3 days, and BMDCs were harvested on day 7.

BMDC were isolated as previously described [17 (link)]. Briefly, femurs and tibias were extracted from 5-week-old Balb/c mice and sterilized by immersion in 70% ethanol for 5 min. The bones were washed thrice with phosphate-buffered saline (PBS; pH 7.4), bone marrow was flushed with complete RPMI medium (Welgene, Gyeongsan, Korea), and red blood cells were lysed. The collected monocytes were resuspended in Iscove’s modified Dulbecco’s medium (Welgene) supplemented with 10% fetal bovine serum (GenDEPOT, Barker, TX, USA), 100 mg/mL streptomycin, 100 units/mL penicillin (Gibco, Carlsbad, CA, USA), 20 ng/mL recombinant mouse granulocyte-macrophage colony-stimulation factor (GenScript, Piscataway, NJ, USA), 20 ng/mL recombinant mouse interlukin-4 (GenScript) and 50 μm β-mercaptoethanol (Sigma-Aldrich, St. Louis, MO, USA). Fresh medium was added on day 3 and BMDC were ready to use on day 7.