An immunoenzymatic system to study in vitro immune responses. Synthesis of antibodies directed toward different -D-galactosidase determinants in lymph node fragment cultures.

A system for studying the antibody response against a single determinant and to all the determinants of a macromolecule (β-D-Galactosidase of ) is described. It consists of culturing fragments of rabbit lymph nodes (either preimmunized or not) and exposing them to antigen . Antibodies secreted into the culture during several days, and up to 3 months in the secondary response, were titrated for: (a) one-hit activation AMEF, the cross-reacting material produced by a point mutant Lac ; and (b) precipitation of wild type enzyme. Titrations of activating and binding antibodies are very sensitive owing to the amplification potential inherent in the enzymatic assays, which allows several antibody measurements on minute samples. In addition antigen decay was followed and correlated with the antibody response, showing faster disappearance when the latter took place. Time-course studies of the antibody response demonstrated that precipitating titres are higher and last longer than activating antibody titres. Repeated challenges showed decay of the memory potential of primed lymph nodes, as well as the possibility of inducing an immune response using non-primed lymph nodes. The results underline the amenability of the present system to the study of primary and secondary immune responses toward restricted portions of a macromolecule.