Distribution of radioiron between internalized ferritin and hemoglobin (Hb) following inhibition of protein degradation. DEC on day 8 of phase II were pulse labeled for 30 minutes with 4 nmol/L⁵⁹Fe-ferritin, then washed and chased with 400 nmol/L unlabeled ferritin. Inhibition of protein degradation was initiated 60 minutes before the ⁵⁹Fe-ferritin pulse by 20 μg/mL leupeptin or 15 μmol/L chloroquine, and inhibitors were present throughout the experiment. Leupeptin, which has a short half-life at 37°C, was added every 12 hours. Equal amounts of protein from cell lysates were analyzed on SDS-PAGE under nonreducing conditions followed by phosphor imaging and scanning with the phosphor imager.⁵⁹Fe-Hb was expressed as percentage of the label in Hb and ferritin. [⁵⁹Fe-Hb/(⁵⁹Fe-Hb +⁵⁹Fe-ferritin)] × 100. Unlabeled Hb was used as a marker (not shown).