Immunological Studies on Guinea Pig Serum and Liver l-Asparaginases: PURIFICATION OF l-ASPARAGINASES BY ANTIBODY PRECIPITATION

Abstract

Abstract Rabbits immunized with partially purified guinea pig serum and guinea pig liver l-asparaginases produced precipitating antibodies to the homologous enzymes. Antisera to guinea pig serum l-asparaginase precipitated the guinea pig liver and agouti serum l-asparaginases but did not cross-react with Escherichia coli B l-asparaginase. Conversely, antisera to guinea pig liver l-asparaginase precipitated the guinea pig serum and agouti serum l-asparaginases but also had no effect on the E. coli B enzyme. The l-asparaginase-antibody precipitates retained varying degrees of the original enzymatic activity, depending on the source of the enzyme as well as on the antiserum used for the precipitation. The ability of guinea pig serum and liver l-asparaginases to cross-react with the heterologous antibodies was utilized to obtain the corresponding heterologous l-asparaginase-antibody precipitates. Separation and reactions of the insoluble l-asparaginase-antibody complexes are described. Immunization of rabbits with l-asparaginase-antibody precipitates increased the production of the l-asparaginase-precipitating antibodies. The resulting specific, secondary antisera were used for the precipitation of the heterologous l-asparaginases. Further enhancement of the antibody titers was observed on injection of the guinea pig serum l-asparaginase-antibody precipitates prepared with the secondary antisera. The insoluble guinea pig l-asparaginase-antibody complexes were solubilized by digestion with papain and purified by DEAE-cellulose column chromatography. The solubilized complexes obtained with the tertiary antisera exhibited a single protein band on polyacrylamide gel electrophoresis and were as effective against Gardner lymphosarcoma as the original guinea pig serum l-asparaginase.