Recombinant murine scf

BMMCs were differentiated and cultured. Bone marrow cells were isolated from femurs of C57BL/6 mice, differentiated in complete RPMI (Gibco), and supplemented with 10% FBS (Gibco), 100 U/mL penicillin/streptomycin (Gibco), 10 ng/mL recombinant murine IL-3 (R&D Systems), and 10 ng/mL recombinant murine SCF (R&D Systems) at 37 °C under 5% CO₂. After 7 days, nonadherent cells were carefully removed and replaced with fresh culture medium to increase the purity of the MCs. This step was repeated every 7 days until adherent cells disappeared (after 5–6 passages). After 5 weeks of culture, BMMC purity was evaluated as the percentage of FcεRIα and c-kit (CD117) cells. BMMCs were used at a purity more than 90%. CD4⁺ T cells were enriched using an isolation kit (Miltenyi Biotech) according to the manufacturer’s instructions. To analyze immune cells infiltrating the colon, collagenase IV (0.05% w/v; Roche) was used to digest the tiny colon tissue (0.1–0.5 cm) for 1 h. Single-cell suspensions were collected and further purified via density gradient centrifugation with 40% (v/v) and 70% (v/v) Percoll-RPMI. LPMCs were collected from the interface and suspended in RPMI medium.^{59 (link)}