Endosomal Trafficking by Actin Filaments Figure 4. Actin pulls endosomes apart for directional movement. HeLa cells internalized Alexa555-EGF for 5 min at 37uC and DMSO or LatB was added subsequently. Nocodazole was added before the addition of the ligands and cells were incubated for 1 h at 37uC. Live cell images were taken at 15 min after internalization using confocal microscopy and a pattern of colors and dots was assigned to each particle. Shape of cells and nucleus are illustrated by dotted lines. Twelve representative tracks are shown for each treatment. The movement of EGF-positive vesicles was tracked with Image J software (A). The movement of the most motile EGF-positive endosomes was tracked relative to the center of the nucleus of each cell. Eight representative tracks (each indicated by a different color) chosen randomly from three independent experiments are shown for each PLoS ONE | www.plosone.org 6 May 2011 | Volume 6 | Issue 5 | e19942