An Accurate Spectrophotom etric Method for Serum Iron and Iron-B inding Capacity without Deproteinization or C entrifugation*!

A simple, direct semi-micro method has been developed for serum iron and latent iron-binding capacity. For total serum iron, into a one cm cuvet are pipetted (a ) 1.5 ml of Na acetate buffer (pH 4.5) containing Na ascorbate and (b ) 0.3 ml of serum. The serum blank absorbance is measured at 562 nm, and 0.2 ml of ferrozine color reagent is added. After incubation (42°, 20 min), the absorbance is again measured. The concentration of serum iron is calcu­ lated by reference to a standard Fe solution (100 fig Fe per dl), with correc­ tion for the absorbance of serum and reagent blanks. For serum latent ircnvbinding capacity, into a one cm cuvet are pipetted (a ) 1.5 ml of tris-HCl buffer (pH 7.8) containing Na ascorbate, (b ) 0.1 ml of serum and (c) 0.2 ml of iron standard solution (400 /¿g Fe per dl). The mixture is incubated (42°, 10 min). The serum blank absorbance is measured at 562 nm, and 0.2 ml of ferrozine color reagent is added. The sample is incubated (42°, 20 min), and the ab­ sorbance is again measured. Correction for the serum blank absorbance is made, and the concentration of unbound iron in the sample is calculated. The amount of added iron minus the amount of unbound iron in the sample is equal to the latent iron-binding capacity. Measurements of serum iron and latent iron-binding capacity by this procedure are comparable in accuracy and superior in precision to measurements by atomic absorption spectrometry.