A research team is tasked with quantifying the protein content of a new drug formulation. They decide to use the Lowry method for protein measurement. As they prepare their samples, they realize that the drug contains reducing agents. How might this influence their protein quantification results using the Lowry method?

Reducing agents can interfere with the Lowry assay because the method depends on copper(II) ions binding to protein and then being reduced by the protein to copper(I), which later reacts with the Folin‑Ciocalteu reagent to produce a blue color. If the sample contains strong reducers, they may reduce copper(II) to copper(I) before it binds to protein, so less copper is available for the protein‑dependent reaction, causing an under‑estimation of protein. At the same time, reducing agents can also reduce the Folin reagent directly, producing a blue color that mimics the protein signal and leads to over‑estimation. For example, a formulation with dithiothreitol could give a high background color and a falsely low protein reading. Therefore, the presence of reducing agents can both suppress the true protein signal and add background noise, compromising the accuracy of the Lowry measurement.