Restricted maturation of antibody-binding characteristics for hapten-specific IgM-plaque-forming cells in mice

Inhibition of plaque formation was used as a method for studying changes in binding characteristics of IgM antibody specific for the hapten, 2,4,6-trinitrophenyl (TNP). By incorporating successive concentrations of TNP ligand into specific hemolytic plaque assays, we were able to obtain inhibition curves and to determine relative ligand binding of anti-TNP antibodies secreted by plaque-forming cell (PFC) populations. The concentration of free hapten required to obtain 50% reduction in IgM PFC decreased with time after immunization, indicating that an increase in average binding affinity of their secreted antibodes occurred. The slopes of the inhibition curves also increased with time after immunization. Maturation with regard to the above two properties was virtually complete for IgM PFC 5–7 days after immunization. The manner by which the slopes of the inhibition curves changed suggested that maturation occurred as the result of a simple selective process, and it appeared to involve primarily precursor cells existing prior to administration of antigen.