Effect of hepcidin on ferroportin and Lamp1 subcellular localization in BMDMs. Presence of ferroportin at the cell surface of macrophages was increased by Fe-NTA (16 hours, 100 μM). In iron-overloaded cells, human hepcidin was added to cell culture media 1 or 3 hours before fixation of the cells for immunofluorescence procedure. Cells were stained with both rabbit antiferroportin (Fpn) and a rat anti-Lamp1 antibody followed by an Alexa 488–conjugated goat antirabbit (green) and an Alexa 588–conjugated goat antirat (red). The merge between the green and the red fluorescence reveals some yellow color corresponding to partial colocalization of ferroportin with Lamp1 protein in hepcidin-treated samples. N indicates the position of the nucleus.