Colocalization of CXCR2 or LDL with EGFP-Rab5 or EGFP-Rab11a in HEK293 cells overexpressing Rab7 or Rab7-T22N.

(A-D) Colocalization of CXCR2 with EGFP-Rab5 or EGFP-Rab11a. HEK293 cells stably coexpressing CXCR2 and Rab7 (A, C) or CXCR2 and Rab7-T22N (B, D) were transiently transfected with plasmids encoding EGFP-Rab5 (A-B) or EGFP-Rab11a (C-D). Cells were treated with CXCL8 (200 ng/mL) at 37°C for 4 hours. Cells were fixed with methanol and incubated with a mouse monoclonal CXCR2 antibody for 30 minutes, followed by a Cy3-conjugated antimouse antibody for 30 minutes. Representative laser-scanning confocal micrographs from 3 independent experiments demonstrating the distribution of CXCR2 (red), EGFP-Rab5, or EGFP-Rab11a (green), and the colocalization (yellow) of CXCR2 with either EGFP-Rab5 (A-B) or EGFP-Rab11a (C-D) are shown. (E-F) Colocalization of LDL with EGFP-Rab5. HEK293 cells stably expressing Rab7 (E) or Rab7-T22N (F) were incubated with Dil-LDL (40 μg/mL) at 37°C for 4 hours, then fixed in methanol. Representative confocal micrographs demonstrating the distribution of LDL (red) and EGFP-Rab5 (green) and the colocalization of LDL with EGFP-Rab5 (yellow) are shown. Images were processed using Photoshop software. Bars, 10 μm.