Approaches to the standardization of serum unsaturated iron-binding capacity

An important aspect in the development and use of serum unsaturated iron-binding capacity (UIBC) procedures is the standardization of reference serum. Spectrophotometric titration of serum transferrin is the oldest of the UIBC methods; however, the literature on this subject indicates that the technique is not being fully exploited and describes a variety of suboptimal conditions. A spectrophotometric titration protocol is described which gives excellent reproducibility with fresh, frozen, lyophilized, and stored serum samples. An indirect UIBC method utilizing differential chelation and ultrafiltration is also described. In this method Fe3+ -NTA (nitrilotriacetic acid), in excess of the anticipated UIBC, is added to a serum sample supplemented with NaHC03. The addition of EDTA (ethelenediaminetetraacetic acid) allows the mobilization of iron from nonspecific macromolecular sites. The ultrafiltrate is analyzed for iron, and the UIBC is calculated. The results were validated via an ultrafiltration titration which was compared with a spectrophotometric titration. The precision of this method is excellent.