CCR2 inhibits in vivo homing of immature B cells. (A-B) Splenocytes (spl) and lymph node (LN) cells from control and CCR2^–/– mice were triple stained with anti-B220 and anti-IgD or anti-IgM. Dot plots show the expression of the markers on B220⁺ cells. Numbers represent percentages of immature population. (A). The graph represents the percentage change in the mature and immature B-cell population in CCR2^–/– mice compared with the control immature and mature populations, which were regarded as 100% from 7 different experiments (B). (C) Splenocytes (spl) and lymph node (LN) cells from control and CCR2^–/– mice were double stained with anti-B220 and anti-CD23. Histograms show CD23 expression on B220⁺ cells. Brackets and numbers show the percentage of CD23^– population. (D) Purification of immature B cells. IgD^– splenocytes from control and CCR2^–/– mice were then separated by anti-B220 magnetic beads as described in “Materials and methods.” Brackets and numbers show the percentage of IgD⁺ and B220⁺ populations. (E-F) Homing of labeled immature B cells to the spleen (E) and LN (F). Labeled immature B cells derived from control and CCR2^–/– mice were injected to control mice. After 3.5 hours, the spleen (E) and LN (F) were collected, and the FITC-positive population was analyzed by FACS. Error bars represent the standard error of the results of all the experiments used to calculate the average.