Two reference measurement procedures are presented here that allow the determination of the iron saturation in human transferrin, based on different molecular properties. The results, directly derived from the number of ions bound to the protein molecule, are traceable to the SI. Up to now, the iron saturation has only been deduced indirectly from the amount-of-substance ratio of serum iron to transferrin in serum. Interlaboratory tests have shown the need for more accurate methods, as the results from many participant test samples for both parameters do not lie within the acceptable range of deviation given by relevant guidelines when different methods or kits are applied. Using isotope dilution, an HPLC ICP-MS procedure was developed in compliance with the requirements of a primary reference measurement procedure. In this manner, the iron saturation was measured with an associated relative expanded measurement uncertainty of 4%. Based on the results, a straightforward Raman procedure was evolved, which allows the determination of the iron saturation in transferrin with an associated relative expanded uncertainty of 7%.
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