File:Dissecting-the-Cell-Entry-Pathway-of-Dengue-Virus-by-Single-Particle-Tracking-in-Living-Cells-ppat.1000244.s003.ogv

English: Endocytic trafficking of DENV. Upon cell entry, the DENV particle (red, surrounded by a white circle) moves rapidly towards an intermediate endosome enriched in Rab5 (blue) and Rab7 (green). This moment is clarified by the appearance of “(A)”, the arrow points at the intermediate endosome. Subsequently, the intermediate endosome carrying the virus particle moves around and merges with other endosomes (indicated by the arrows): a Rab5-positive endosome (B) and an intermediate endosome (C). Next, the intermediate endosome carrying the virus matures into a late endosome through a gradual disappearance of Rab5 (D). Membrane fusion occurs from within the Rab7-positive endosome (E). Snapshots of this video are shown in Figure 3B . During real-time imaging, DiD-labeled DENV particles were recorded at 2 Hz, whereas the excitation of Rab5-eCFP and Rab7-eYFP were alternated at 0.5 Hz. Due to this recording scheme, the CFP and YFP frames are updated every 4 frames in the video, whereas the DiD signal is updated every frame, which might give the impression that the endosomal markers are lagging behind the virus particle. The hallmarks of colocalization are that the DiD spot and the Rab signal at least show partial overlap when they are simultaneously excited, and that both signals are linked and move together in time. The playback speed is 10× real-time.