In the recent literature an inverse relationship between iron status and serum aluminum levels has repeatedly been reported in dialysis patients. To check whether this observation is, at least in part, due to an interference of iron with the protein binding of aluminum, we studied the effect of the latter element on both the number of free binding sites on transferrin (Tf) and on the affinity of the protein for aluminum. For the purpose of this, a recently developed HPLC-ETAAS hybrid method was used, allowing protein-binding studies at clinical relevant metal concentrations and under contamination-free conditions. After we incubated apo-Tf with iron and aluminum which were added in amounts equivalent to the calculated number of metal-binding sites on the protein (i.e., 2 mol metal/mol Tf), we found that Tf can be saturated for 100% with iron. However, for aluminum only a 23% aluminum-Tf saturation was observed. In Tf solutions with iron saturations ranging between 0 and 60% as well as in the serum of 15 subjects with iron-Tf saturations varying between 12 and 48%, a significant (p < 0.001) negative correlation between the degree of iron-Tf saturation and the percentage of aluminum (added in amounts equivalent to the number of the remaining binding sites on Tf) bound to Tf was noted (y = -0.26x + 24.5, r = -0.87 in serum). It is concluded that the iron-Tf saturation influences the Tf binding of aluminum not only by occupying binding sites otherwise available for aluminum, but also by lowering the affinity of Tf for aluminum. The effects of iron on serum aluminum levels and bone aluminum deposition are discussed.