Loss of VPS45 leads to a defective early-to-late endosome maturation. Nonmodified control and VPS45 KO HeLa clones were transduced with recombinant lentiviral vectors to coexpress mCherry-Rab5 and mCerulean-Rab7. Dynamic changes of Rab5-positive and Rab7-positive vesicles were imaged during the uptake and transport of fluorescent cargo (microspheres-633). Arrows point to microspheres. (A) Representative images of cargo trafficking in single vesicles of control and VPS45 KO clones are shown. Scale bars, 1 µm. (B) The corresponding total fluorescence traces of Rab5 (red) and Rab7 (green) over time averaged from 5 independent experiments are shown. For visualization purposes, only every second frame is shown for the first 12 minutes in VPS45 KO clones. Error bars indicate mean ± standard error of the mean.