A specialized serum ferritin assay has been developed for the detection of iron deficiency in epidemiologic studies. An enzyme immunoassay (EIA) was employed to eliminate the need for radioisotopes. The problem of low sensitivity inherent with the EIA for serum ferritin was eliminated by the use of monoclonal immunologic reagents. The working range of the assay is 1-100 micrograms/L with a sensitivity of 0.5 micrograms/L. Excellent agreement in serum ferritin levels was observed between the present method and the two-site immunoradiometric assay (IRMA), while the variability at low ferritin concentrations was significantly less with the EIA. Because only 10 microliter of serum is required for each assay, duplicate measurements can be performed on a single capillary tube of blood. When an automatic microtiter plate reader for optical density measurements is used, 80-100 duplicate determinations can be completed by one technologist in a single working day.

This study was undertaken to assess the relationship between iron absorption and the concentration of duodenal iron proteins in normal subjects and patients with idiopathic hemochromatosis (IH). Biopsies were obtained endoscopically from the duodenum in 17 normal subjects, 3 of whom were mildly iron deficient, and 7 patients with untreated IH. The absorption of both heme and nonheme iron was increased in IH despite a 20-fold elevation in serum ferritin. Immunoassays using MAb were used to measure transferrin, H-rich ferritin, and L-rich ferritin in mucosal samples. Mucosal transferrin concentrations in normal subjects did not correlate with either iron status or iron absorption, indicating that mucosal transferrin plays no physiological role in iron absorption. Mucosal transferrin was significantly lower in IH, presumably because of a decrease in mucosal transferrin receptors. Mucosal H and L ferritin concentrations were directly related to body iron stores and inversely related to iron absorption in normal subjects. In IFH, mucosal H and L ferritin failed to increase in parallel with the serum ferritin, but were appropriate for the level of iron absorption. The relationship of mucosal H/L ferritin in IH did not differ from that observed in normal subjects. Our findings indicate that the major abnormality in duodenal iron proteins in IH is a parallel decrease in the concentration of H-and L-rich ferritin. It is-not evident whether this is the result or the cause of the absorptive abnormality.

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Human spleen ferritin was labelled with 131I and injected into two normal men. The labelled ferritin left the plasma rapidly. The experimental clearance curve could be fitted with accuracy into two single exponential functions. The first component, T 1/2 = 9 min, accounted for the clearance of about 90% of the labelled ferritin. Surface counting showed uptake of 131I by the liver but not by the spleen. Such a rapid plasma turnover is similar to that found after injection of tissue ferritins into experimental animals but contrasts with the slow turnover previously found for 131I-labelled human plasma ferritin. Differential clearance of isoferritins from the plasma is an important factor explaining the biochemical and immunological differences between tissue and plasma ferritins.

We have used undifferentiated human promyelocytic HL60 cells to study the binding of radioiodinated human ferritin in vitro. Specific binding of human heart ferritin could be demonstrated at 37 degrees C, whereas no binding of liver ferritin could be found. The uptake of labelled heart ferritin was abolished by incubation at 4 degrees C, by prior treatment of the HL60 cells with pronase and by the addition of human plasma to the medium. On the other hand, the addition of excess unlabelled human liver or rat liver ferritin had no effect on the uptake of labelled human heart ferritin. Dissociation studies showed that about 55% of the bound heart ferritin radioactivity could be released by incubation with medium alone and at least 90% with excess unlabelled heart ferritin. Over 70% of the dissociated ferritin could be precipitated with polyclonal anti-ferritin serum or trichloroacetic acid. More than two-thirds of the radioactivity which could not be released after washing in medium alone was recovered in the soluble intracellular fraction following cell lysis. Almost all of the soluble radioactivity could be precipitated with the polyclonal antiserum, indicating that very little lysosomal degradation of internalized heart ferritin had occurred. The present studies demonstrate a protein-mediated binding mechanism for acidic isoferritins on HL60 cells. These observations agree with published evidence that ferritin is often associated with cell membranes and are consistent with a possible role for the protein in the regulation of haematopoiesis or in iron transfer.