Fig. 6. Imaging of EGF and EEA1. (A) COS-7 cells expressing GFP-EEA1 (green) were exposed to 50 ng/ml Alexa-Fluor-568-EGF (red) and imaged for the times after ligand addition, as indicated in each panel. After 3 minutes cells were washed and imaged by TIRF-M with an incident angle set to visualize 300 nm from the coverslip. Colocalized voxels are depicted in white, and indicated with arrowheads. Arrows point to the appearance of a ring-like structure containing EEA1, to which EGF-containing vesicles appear to attach. (B) Higher resolution series depicting the association of EGF-containing vesicles (arrow) with EEA1-enriched endosomes. (C) Optical sections at 250 nm intervals through the entire cell volume were projected into a single 2D image.