Chemokine binding and signaling properties of D6 on peritoneal cavity B1 cells. (A) CCL2^AF647 uptake by WT and D6-deficient (KO) peritoneal cavity B1a/b (CD19⁺CD11b⁺) or T cells (CD19⁻CD11b⁻CD5⁺) in the presence of a 10-fold molar excess of the indicated unlabeled chemokines. No comp indicates CCL2^AF647 uptake in the absence of competitor. The average mean fluorescence intensity (MFI) of competed samples is shown as a percentage ± SEM of the average MFI of uncompeted WT uptake. Asterisks mark chemokines that significantly reduced CCL2^AF647 uptake by WT or D6-deficient KO cells (P < .01). (B) Gates used to gauge Ca²⁺ fluxes in B1a/b and macrophages. (C-D) Representative Ca²⁺ flux profiles of Fluo-4/Fura-Red–loaded WT and D6-deficient KO peritoneal cavity B1a/b cells (C) and macrophages (D). The points of chemokine (50-100nM) and ionomycin addition are indicated with arrows. Cells in the far right panel of panel C were incubated in 100nM CCL22 for 1 hour at 37°C before analysis. (E) Migration of WT B cells from peritoneal cavity lavages to CCL22 or CXCL13. Migration in medium alone is indicated by the unfilled square. The mean number of live cells ± SEM migrated from 4 replicate wells is presented as a percentage of input. All data are representative of 3 or more independent experiments.