Generation of Ig-isotype diversity was studied in hemolytic plaque-forming cells (PFC) of mice immunized with sheep erythrocytes (SRBC). Spleen cells were taken at various days after immunization, cultured in vitro for 24 hr in the presence of bacterial lipopolysaccharide (LPS), and PFC occurring in these mass cultures were micromanipulated and individually cultured for 48 hr. Daughter PFC were revealed by direct or indirect procedures in order to study the Ig-isotype they produced.
Parental PFC of all three Ig isotypes studied, IgM, IgG, and IgA, generated daughter PFC, producing the same isotype but with maximum yields on different days: postimmunization day 3 for IgM PFC, day 4 for IgG PFC, and day 5 for IgA PFC. A proportion, but not all, of parental IgM PFC generated daughter IgG producers only during the first 3 days after immunization and IgA producers during the 4th to 6th post-immunization days. Parental, especially 4th day IgG PFC also generated daughter IgA producers, thus showing that IgA producers could stem from both IgM and IgG precursors in addition to IgA-producing precursors. It was also found that IgG PFC could present a reverse switching and generate IgM producers. This suggests that the overall orderly IgM, IgG, IgA conversion is possibly not strictly programmed by an internal control of B cells but submitted to some extent to microenvironmental influences.
This work was supported by CRL-77 5 070 I Grant of I.N.S.E.R.M. and LA 143 Grant of C.N.R.S.