Fig. S6.

Colocalization of internalized Tf with TfR-GFP on axonal retrograde carriers. (A) Rat hippocampal neurons were cotransfected at DIV5 with plasmids encoding TfR-GFP and mCherry-tubulin and were incubated at DIV8 with Alexa 647-conjugated transferrin (Tf-Alexa 647) for 30 min at 37 °C. After washing with PBS, cells were analyzed by live-cell imaging. TfR-GFP, Tf-Alexa 647, mCherry-tubulin, and merged images are shown in A. Arrows indicate the position of the AIS. (Scale bar: 10 μm.) (B) Magnifications of the boxed regions in A. The top upper strips show single frames, and the two middle panels show negative grayscale kymographs for up to 200 s of imaging from Movie S4. The bottom panel shows the merged trajectories of carriers having both TfR-GFP and Tf-Alexa 647 (yellow lines), only TfR-GFP (green lines), or only Tf-Alexa 647 (red lines). Lines with negative and positive slopes represent carriers moving in anterograde and retrograde directions, respectively. Vertical lines represent stationary foci.