Peripheral blood mononuclear cell‐derived human mast cells
Human peripheral blood mononuclear cell‐derived mast cells were generated as previously described by Folkerts et al. 33 Briefly, peripheral blood mononuclear cells were obtained from buffy coats of healthy blood donors and CD34+ precursor cells were isolated using the EasySep Human CD34 Positive Selection Kit (STEMCELL Technologies). CD34+ cells were maintained for 4 weeks under serum‐free conditions using StemSpan medium (STEMCELL Technologies) supplemented with recombinant human IL‐6 (50 ng/mL; Peprotech), human IL‐3 (10 ng/mL; Peprotech), and human Stem Cell Factor (100 ng/mL Peprotech). Thereafter, the cells were maintained in IMDM Glutamax I containing sodium pyruvate, supplemented with 0.1% 2‐mercaptoethanol, 0.5% BSA, 1% insulin‐ 175 transferrin selenium (all from Invitrogen), ciprofloxacin (10 µg/mL; Sigma‐Aldrich), IL‐6 (50 ng/mL; Peprotech), and human Stem Cell Factor (100 ng/mL; Peprotech). After 8‐12 weeks, PBCMCs were tested for maturity by Giemsa or toluidine blue staining and beta‐hexosaminidase release assays.