Tf-endosomal motility is reduced upon interaction with mitochondria. MitoTracker orange–labeled MDCK-PTR cells were pulsed with AF488-hTf for 2 min at 37°C and chased, and time-lapse images of 15-s duration (139-ms interval) were acquired at 5 min after the hTf pulse. (A) Double-labeled fluorescence image from a representative time lapse (a); corresponding 3D render where AF488 hTf-endosomes are green spots and mitochondria are red surfaces (b); 3D-rendered image subjected to a DT 0–based kiss and run filter selecting only those Tf-endosomes that are in close proximity to mitochondria (c; see Video 5); magnified ROI (d; white box in c); magnified ROI (white box in d) where the selected hTf-endosome (yellow spot) is in close proximity to a mitochondrion (e). Bars: (a–d) 10.0 µm; (e) 1.0 µm. (B) Videomicrograph (top row, fluorescence; bottom row, 3D rendered) of the kiss and run interaction between the selected hTf-endosome and mitochondrion (A e) proceeds from left to right at a 139-ms frame interval. Graph represents the hTf-endosomal instantaneous speed (blue line) with respect to its corresponding DT-based distance from the mitochondrion (red line) over time. Dashed box in videomicrograph and graph indicates the kiss phase. hTf-endosomal instantaneous speed is markedly reduced when it is in close proximity to the mitochondrion, denoted by the overlapping of the red line with the x axis. See Videos 6 and 7, which show two additional hTf-endosomal kiss and run events. Bar, 0.5 µm.