SDF-1 induced internalization of CXCR4 from surface of early and mature B cells. Total bone marrow lymphocytes were incubated for 30 minutes at 37°C with 125 nmol/L of SDF-1 and placed on ice immediately after incubation to prevent resurfacing of the receptor. Cells were subsequently washed in ice-cold glycine buffer to remove cell bound SDF-1. Next, the cells were stained for CXCR4 expression in parallel with surface markers defining (A) early and (B) late lineage B cells (see Fig 1A). The early lineage B-cell population contained both pro-B and pre-B cells, whereas late lineage B cells contained immature and mature B cells. The shaded part of the histogram represents untreated cells; the gray line represents cells incubated with SDF-1; the dashed black line represents antibody isotype control. The results are representative of 3 experiments with primary B cells.