CCL19- and CCL21-induced calcium flux. (A) Purified unstimulated eosinophils and IL-5–primed eosinophils were loaded with fluo-4 AM, and calcium flux was recorded using flow cytometry. Baseline was established for 35 seconds before chemokines were added. Arrows indicate time of addition of CCL19 (1 μg/ml), CCL21 (1 μg/ml), or CCL11 (100 ng/ml). Data are representative of four separate experiments from separate donors. (B) Cross-desensitization of CCL19 and CCL21 for calcium flux. Calcium flux of IL-5–primed eosinophils was measured as above. Arrows indicate time of addition of CCL19 (5 μg/ml) or CCL21 (5 μg/ml). Data are representative of three separate experiments from separate donors. (C) In the upper panel, IL-5–primed eosinophils were pretreated with pertussis toxin (PTX) (1 μg/ml). In the lower panel, eosinophils were resuspended in calcium-free medium with 2 mM EGTA for 10 minutes before the addition of CCL19 and CCL21. Arrows indicate time of addition of CCL19 (1 μg/ml) or CCL21 (1 μg/ml). Data are representative of three separate experiments from separate donors.