If the protein concentration in a sample is underestimated when using colorimetric assays, what is the most likely underlying cause of this effect?

When a colorimetric assay underestimates protein, the most common reason is that a component in the sample interferes with the dye–protein interaction or the reaction chemistry. For example, detergents such as SDS or strong reducing agents can prevent the dye from binding to the protein, so the measured absorbance is lower than it should be. Because the assay relies on a specific color change, anything that blocks the dye’s access to the protein or quenches the color will make the protein look less concentrated. A simple illustration is a Bradford test performed on a lysate containing SDS; the dye cannot bind, so the absorbance is low and the protein level is underestimated.