Much evidence supports the thesis that secreted proteins are made on membrane-attached ribosomes, whereas proteins retained within cells are generally synthesized by ribosomes lying free in the cytoplasm [1 ]. Ferritin, a retained protein found in many tis- sues, consists of an outer shell of apoferritin subunits surrounding a core of iron salts. Studies in our labo- ratory [2] and by Redman [3] agree in showing pre- ferential synthesis of apoferritin by the free ribosomes of rat liver; after incubation with labeled amino acids, free ribosomes show many more nascent chains of apoferritin than total or membrane-attached ribosomes, whereas the reverse is true for albumin. Nevertheless, Redman observed that, a few minutes after intact rats were injected with [14C]leucine, only 75% of the radioactivity in the nascent apoferritin of the liver was recovered in the free ribosome population, the re- maining 25% being associated with membrane-bound ribosomes. Contamination with free ribosomes could account for about 10% of this. Puro and Richter [4] also used in vivo injection of labeled leucine and found that labelwas incorporated into nascent liver apofer- ritin about 20-30% as well by microsomal ribosomes as by free ribosomes. Contamination of their micro- some fraction with free ribosomes did not exceed 4--8%. While these studies indicate preferential syn- thesis of ferritin on free liver ribosomes, they leave doubts about these as the exclusive site of synthesis. The size of polysomes synthesizing a given protein depends on the number of amino acid residues for which the messenger codes. Although horse spleen apoferritin appears to have 24 identical subunits with a mol. wt. of 18 500, we [5] have recently observed by gel electrophoresis that rat liver ferritin contains two types of subunits with molecular weights of 18 500 and 12 000 respectively. We have therefore examined the free and membrane-bound ribosomes of rat liver for the presence of polysomes consistent with synthesis of these two subunits by reacting each population of ribosomes with ferritin antibody coupled to 125 I. On free polysomes, the antibody was bound by trimers and tetramers, consistent with a peptide, having a mol. wt. of 12 000, whereas the ferritin antibody attached to pentamers in the mem- brane-bound ribosome population, consistent with a product of 17 000 molecular weight. We conclude that free and membrane-bound ribosomes contribute different subunits to liver ferritin. 2. Materials and methods 125 Iodine (sodium salt, carrier free) was obtained from New England Nuclear. Lactoperoxidase was purchased from Calbiochem., and heparin from Nu- tritional Biochemicals. Other chemicals were reagent grade. Rabbit antiserum against horse spleen ferritin was generously supplied by Dr. Linder (M.I.T.). Most of the media were made up in TKM buffer (50 mM Tris-HC1, pH 7.8; 25 mM KC1; 5 mM MgCI2) contain- ing heparin to inhibit ribonuclease.