Ferritin-iron cores are present in exosomes. (A) RAW264.7 macrophages were incubated for 24 hours in a mixture of OptiMEM I medium and Dulbecco’s modified Eagle medium (1:1 volume ratio) supplemented with 10% heat-inactivated fetal bovine serum, 2 mM l-glutamine, 100 U/mL penicillin, 100 μg/mL streptomycin, 1 mg/L BSA, 20 mM β-mercaptoethanol, and 100 μg/mL FAC. To precipitate exosomes, cells were harvested, and medium was collected and centrifuged at 100 000g for 1.5 hours. Samples were then separated by SDS-PAGE and analyzed by western blot with anti-ferritin L-subunit and anti-TSG101 (serving as exosomal marker) antibodies. The results of 1 out of 4 experiments are shown. (B) Exosomal samples were resuspended in 0.1% Glutaraldehyde, and a drop was mounted on an ion-coated copper grid supported by a carbon-coated film. The sample was stained with 1% uranyl acetate and visualized by TEM. Scale bar represents 100 nm. (C) Exosomal samples were captured by cryo-TEM. Vitrified unstained specimens were loaded to a Tecnai T12 G2, operating at 120 kV, and examined at a low dose to minimize radiation damage. The arrows point to iron cores. Scale bars represent 100 nm.