Iron accumulation and increased ferritin expression in the duodenum of IRP2-deficient mice. (A) The proximal part of the duodenum from 10-week +/+ and -/- males was analyzed by Prussian blue staining (top panels) or by immunostaining with anti-ferritin L-chain (middle panels) or anti-ferritin H-chain antibodies (bottom panels). Prussian blue coloration was counterstained with Nuclear Fast Red and ferritin immunostaining with hematoxylin. Pictures were acquired on a Axiophot microscope (Zeiss, Jena, Germany) with a × 20 objective. (B) Expression of ferritin H- and ferritin L-chains in groups of 4 +/+, +/-, and -/- 10-week-old males. Total protein (40 μg) was resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and subjected to Western blot analysis (top panels). Ferritin H- and L-chain mRNA levels were analyzed by Northern blotting using 10 μg total RNA from the same mice (bottom panels) and β-actin mRNA as a standard. Western blot and Northern blot signals were quantified and are presented as a histogram (bottom) after normalization for β-actin expression. Error bars indicate standard deviation. In Western blots, the ferritin L-subunit resolves into 2 bands, the bottom one corresponding to a cleavage product. Both bands were taken into account for quantification. IRP2 expression was below the detection limit.