Elimination of turbidity interference in serum iron colorimetric assay by enzymatic proteolysis

Abstract

We describe a modification in the commercial colorimetric method for the determination of serum iron by using Ferrozine® . The modification was proposed because during the conventional procedure, turbidity observed when the serum of animals submitted to surgery was used interfered with the assay. We added to the original method, a previous treatment of the serum with proteolytic enzymes. This modification was also tested using plasma samples, although this was not recommended when the original method was used. The results demonstrated that: a) the treatment with a mixture of trypsin and chymotrypsin was effective in order to eliminate turbidity; b) there was no difference between the standard curves obtained by the conventional and the modified method for control assays; c) the absorbencies of the samples of serum and plasma submitted to proteolysis, estimated by the addition of different concentrations of iron, were directly proportional to iron concentrations; d) the pre-treatment with enzymes allowed the utilization of plasma; e) the pre-treatment with guanidine. HCl was not effective.

Keywords

• Iron assay
• enzymatic proteolysis
• interference