Gene family use and somatic mutation in primary and secondary fluorescein-specific IgM antibody responses.

A comparative analysis of the DNA sequences of primary and secondary IgM, fluorescein-specific antibodies was performed. These antibodies were secreted by hybridomas generated following fusion of immunized BALB/c mouse lymphocytes and SP2/0 myeloma cells. Our results show that primary and secondary fluorescein-specific IgM antibodies use a variety of segments from the variable region of the immunoglobulin heavy chain locus (VH), with members of the J558 and 7183 VH gene families predominating in both populations. D regions from the DF116 and DSP2 families were used exclusively in our primary antibody sample and predominated in the secondary response. In the primary antibodies, 15 out of 18 definable D regions were transcribed in reading frame one, but in the secondary antibodies the three reading frames were used stochastically. Secondary IgM antibodies showed a higher frequency of somatic mutation than their primary counterparts, but we could detect no evidence of selection for mutations in the complementarity determining regions as compared with the framework regions. It appears that fusion of secondary cells, 3-6 days after immunization, is able to 'capture' the IgM-producing population of B cells at a stage in their development following mutation but prior to antigenic selection.