Monoclonal and Polyclonal Antibodies Against Porcine Mitochondrial Aspartate Aminotransferase: Their Inhibition Modes and Application to Enzyme Immunoassay

Monoclonal and polyclonal antibodies against porcine mitochondrial aspartate aminotransferase (m-AST) were prepared in order to study their effect on the kinetics of the enzyme and their possible use as diagnostic reagents. The most stable hybridoma clone, designated MH-1, was selected and cultured for mass production of the monoclonal antibody MA-1. MA-1 was purified by affinity chromatography with m-AST as a ligand. The m-AST activity was inhibited uncompetitively by preincubation with MA-1, but preincubation with the polyclonal antibody uncompetitively by preincubation with MA-1, but preincubation with the polyclonal antibody raised in a rabbit resulted in noncompetitive inhibition of the enzyme. These results suggest the usefulness of a monoclonal antibody for studying the mechanism of catalysis. Sandwich enzyme immunoassay methods for m-AST using both polyclonal and monoclonal antibody-coated polystyrene balls were established and permitted the determination of porcine m-AST on the order of 10(-11) and 10(-10)M, respectively.