Competitive binding assays for high-affinity binders in the presence of endogenous ligands: Application to biotin-binding proteins
✍ Scribed by Richard W. Schreiber Jr.; Michael A. Letavic; Timothy J. McGahan; Harold B. White III
- Publisher
- Elsevier Science
- Year
- 1991
- Tongue
- English
- Weight
- 731 KB
- Volume
- 192
- Category
- Article
- ISSN
- 0003-2697
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✦ Synopsis
Endogenous ligands complicate radioligand-binding assays of high-affinity binding proteins by obscuring binding sites or by diluting the labeled ligand. We have developed a mathematical model for such systems where radioligand and endogenous ligand are structurally identical. Data which relate radioligand binding at equilibrium as a function of sample volume can be plotted such that the concentrations of endogenous ligand and binder are graphically determined; however, a more precise determination may be done by nonlinear regression with the aid of a microcomputer. The method is demonstrated for the assay of biotin-binding proteins in the presence of a range of endogenous biotin concentrations below and above that required to saturate the binding sites.