The generation of antibodies capable of catalyzing the unassisted hydrolysis of unactivated amides has been an enduring goal of research in the field. Antidialkylphosphinate 1 monoclonal antibodies were screened for their ability to catalyze the hydrolysis of four methyl esters and four primary amides at pH 5.0, 7.0, and 9.0. One of 68 antibodies, 13D11, enantiospecifically hydrolyzed the C-terminal carboxamide of a dansyl-alkylated derivative of (R)-phenylalaninamide (2b). At pH 9.0, 13D11 catalyzed amide hydrolysis with a k(cat) of 1.65 X 10(-7) s(-1) and a K-m of 432 mu M and was stereospecifically inhibited by hapten with a K-i of 14.0 mu M. A shorter, acetylated derivative 3b was not hydrolyzed by 13D11, demonstrating that dansyl-alkyl binding interactions are essential for catalysis. Equally active antibody preparations were obtained from two separate batches of ascites. The antibody Fab’ fragment was prepared, purified, and found to retain full activity. Amidolytic activity was not abolished by any of nine inhibitors of natural proteolytic enzymes. The rate of uncatalyzed amide hydrolysis was experimentally determined to be 1.25 x 10(-9) s(-1), indicating an antibody catalytic rate enhancement (k(cat)/k(uncat)) of 132.