What is the first step in the quantitation of serum iron?

• A. Direct reaction with appropriate chromogen
• B. Iron saturation of transferrin
• C. Free iron precipitation
• D. Separation of iron from transferrin

Answer: (D)

Knowing that most serum iron is bound to transferrin, the first move in quantitation is to separate iron from this binding protein to release the iron so it can be measured. This initial step usually uses acid or another method to denature transferrin and free Fe ions. Once iron is liberated, it’s reduced to the Fe2+ form and then forms a colored complex with a chromogen that is read spectrophotometrically. So separation is the essential prerequisite that makes the subsequent reduction and chromogen reaction possible. Direct reaction with the chromogen can’t occur until iron is free, and iron saturation describes a binding state rather than a procedural step, while precipitating free iron would come after iron has been released.