Herein is reported a method for determining in a sample the (total, i.e. binding competent) amount of a ligand of a ligand-binding protein (therapeutic) comprising the following steps in the following order: subjecting the sample to an acid treatment, forming in solution a non-covalent complex comprising i) an anti-ligand antibody, ii) the ligand, and iii) labelled ligand-binding protein, by adding anti-ligand antibody and labelled ligand-binding protein to the sample, and determining the amount of the complex and thereby determining the amount of the ligand of the ligand-binding protein.