The Folin-Ciocalteu reagent can be used effectively for measuring protein concentrations in all types of biological samples without any limitations.

The Folin‑Ciocalteu reagent is designed to detect phenolic and other reducing groups, not the peptide bonds that make up proteins, so it does not give an accurate measure of protein concentration. When used with proteins, the reagent reacts only with certain amino acids such as tyrosine or tryptophan, producing a color change that depends on their number rather than on total protein mass. Because many biological samples contain sugars, nucleic acids, or detergents that also reduce the reagent, the resulting absorbance can be heavily skewed, leading to over‑ or under‑estimation of protein levels. For example, a plasma sample that contains a high amount of bilirubin will give a much higher signal than a protein‑free sample, even though the protein content is the same. Therefore, Folin‑Ciocalteu is not a reliable universal method for protein quantification and should be replaced by assays such as Bradford, BCA, or Lowry that are specifically calibrated for proteins.