Studies on lymphokines: the production of antibody to guinea pig lymphotoxin and its use to distinguish lymphotoxin from migration inhibitory factor and mitogenic factor.

Guinea pig lymphotoxin, which was produced by stimulating immune lymph node cells with the specific antigen, ovalbumin, was partially purified by the sequential use of Pevikon block electrophoresis, ion exchange chromatography on DEAE-cellulose, and polyacrylamide disc gel electrophoresis. The lymphotoxin recovered from the final step was purified approximately 200- to 300-fold as compared to the starting material, but nevertheless contained at least one major contaminant protein. Rabbits were immunized with the partially purified lymphotoxin, and an antiserum capable of neutralizing the cytotoxic effect of lymphotoxin was obtained. The anti-lymphotoxin activity was shown to be due predominantly to IgM immunoglobulin. The antiserum neutralized concanavalin A-induced lymphotoxin with equal efficiency as antigen-induced lymphotoxin. On the other hand, the anti-lymphotoxin serum did not neutralize preformed mitogenic factor or migration inhibitory factor (MIF) which had been produced by antigen-stimulated immune lymph node cells. Thus these activities appear to be due to substances which are antigenically distinct from lymphotoxin. Likewise, anti-lymphotoxin serum, when present in cultures of antigen-stimulated immune lymphoid cells, caused no diminution in the amount of mitogenic factor or MIF which was produced, indicating that it had no direct deleterious effect on the function of the lymphoid cells responsible for producing these lymphokines.