Cell isolation, culture, and handling

RBL-2H3 cells (American Type Culture Collection CRL-2256) were cultured at 37°C and 5% CO₂ in Eagle’s minimum essential medium (MEM; Sigma-Aldrich) supplemented with 15% FCS, 4 mM l-glutamine, and 1% penicillin-streptomycin-neomycin solution (Sigma-Aldrich). Twenty-four hours before imaging (or for experiments requiring suspension culture), cells were seeded at 1 × 10⁶/ml and incubated in 50-ml culture tubes overnight on an end-over-end rotator (6 rpm) at 37°C. Primary human basophils were isolated from leukocyte cones provided by UK National Health Service Blood and Transplant. Nucleated cells were separated from erythrocytes with the HetaSep aggregation agent (STEMCELL Technologies) as per the manufacturer’s instructions. Basophils were isolated through negative selection with the EasySep Human Basophil Enrichment Kit (STEMCELL Technologies) and used immediately after isolation.