Expression and subcellular localization of ferroportin after erythrophagocytosis. (A) BMDMs were treated with artificially aged red blood cells as described in “Materials and methods.” After 8 hours, membrane preparations and cytosolic extracts were isolated and processed for Western blot analysis using antiferroportin (top, membrane fraction) or anti–H ferritin (bottom, cytosolic fraction). Middle panel shows Red Ponceau (RP) staining of PVDF membrane after transfer used as a control of loading for the ferroportin detection. The position and size in kilodaltons of molecular mass markers are indicated on the right. (B) Immunofluorescence staining of endogenous ferroportin in control cells or in cells 8 hours after erythrophagocytosis. After EP, ferroportin presents both strong vesicular and cell-surface staining (arrows).