A part of the pool of AQP-1 colocalizes with the TfR and is found in multivesicular endosomes in reticulocytes. After purification by Percoll gradient, reticulocytes (A-B) and red blood cells (C) were fixed with acrolein and deposited on cover slips pretreated with poly-l-lysine. After permeabilization (Triton X-100), the cells were immunostained for GPA (green) and AQP-1 (red) (A,C), or TfR (green) and AQP-1 (red) (B). The slides were examined in a confocal microscope (Zeiss LSM 510 META camera, Zeiss Plan Apochromat 100×/1.4 NA oil objective, scan zoom 2.0) and pictures acquired and processed with Zeiss Laser Scanning Microscope LSM 510 software, Version 3.2 SP2. For visualization of TfR and AQP-1 in multivesicular endosomes and exosomes, reticulocytes were fixed with a mixture of paraformaldehyde, glutaraldehyde, and sucrose. After dehydration and embedding, thin sections of cells were mounted on nickel grids, blocked, and then immunostained for the TfR (D) or AQP-1 (E-F). Sections were examined in a Philips-410 electron microscope equipped with AMT XR41 Biological 2k side-mounted CCD camera, and images were acquired with AMT600 software and analyzed with Photoshop CS (Adobe Systems). After treatment with dynasore 80 μM or the solvent, DMSO, alone for 20 hours (G), 10⁶ cells (left) and exosomes, resuspended in the same volume of Laemmli buffer,²³  (right) were examined by SDS-PAGE and immunoblotting for the proteins indicated.