Restriction in IgM expression--VI. Affinity analysis of monoclonal anti-dansyl antibodies.

Affinity restriction in the IgM response to the 5-dimethylaminonaphthalene-1-sulfonyl (dansyl) group was studied with 56 monoclonal IgG and IgM affinity-purified antibodies. These were generated by immunization with dansyl-Ficoll or dansyl-B gamma G. Association constants for N epsilon-dansyl-lysine were determined by fluorescence titration based on resonance energy transfer. The antibodies were also characterized with respect to the emission maximum of the antibody-ligand complex. The distribution of binding constants covered a 100,000-fold range, demonstrating thereby a restriction in IgM affinity relative to IgG affinity. In the secondary response the IgG affinity exhibited a ceiling of about 1 X 10(10) M-1, a value in accord with the size and hydrophobic nature of the dansyl group. In the case of IgM, with one exception, the value of 1 X 10(7) M-1 was not exceeded with IgM antibodies isolated from 28 hybridomas. This system, therefore, provides further evidence for the restriction in expression of IgM as a general feature of the immune response.