Fig. 7. CXCR4 levels, recycling and migration. (A) Western blot analysis of CXCR4 protein levels in TAM2D2 cells and C2B transfectants. After CXCR4 staining, blots were stripped and reprobed with actin antibody. (B), AMFR (autocrine motility factor receptor), IGF1R (insulin-like growth factor 1 receptor) and LFA-1 (aLb2 integrin) in TAM2D2 cells and C2B transfectants. (C) Cell surface levels of CXCR4 in TAM2D2 cells and C2B and C2B/CXCR4-GFP transfectants. (D) Migration towards 100 ng/ml CXCL12. Results are given as the mean ± s.e.m. of the migration index, which is set to 1 for the control cells in each individual experiment. Shown are averages of three experiments. (E) Recycling of CXCR4 in TAM2D2 cells and C2B transfectants. Cells were stained with CXCR4 antibody before or after 2 hours treatment with 100 ng/ml CXCL12; or after CXCL12 treatment and subsequent overnight recovery in medium without CXCL12, supplemented with cycloheximide. The experiment was repeated three times; representative results of one experiment are shown. (F) CXCR4 levels in TAM2D2 cells, assessed by western blotting at 2 and 4 hours in the presence of cycloheximide, after treatment with 100 ng/ml CXCL12, in both TAM2D2 cells and C2B transfectants. Staining with actin antibody was used as loading control.