DTPA-coupled proteins—procedures and precautions

Abstract

While demonstrating that the cyclic anhydride of diethylenetriaminepentaacetic acid (DTPA) may be used to covalently attach this chelator to proteins, it was necessary to develop several new procedures and techniques. We were able to show that coupled proteins may be labeled with indium-111 ( 111In) simply by transcomplexation from a weaker complex such as the acetate to avoid subjecting the protein, even momentarily, to the acidity of the chloride (however, we have also observed that too high a concentration of acetate and other complexing agents such as citrate will interfere with protein labeling by competing with DTPA for the radioactivity). In agreement with other investigators using bifunctional chelate methodology we have observed that trace metals may interfere with the labeling. Accordingly, we developed a simple paper Chromatographic assay which may be used to establish whether trace metals are present at interfering concentrations and to identify their sources. We have also employed a hydrolyzed control assay to determine whether and to what degree the radioactivity is bound to protein other than at the DTPA groups. We have developed a simple method of measuring the average number of DTPA groups per protein molecule which involves labeling the products of coupling with 111In. Procedures used and precautions observed in the radiolabeling of proteins by these methods are discussed.