Abstract
An in vitro splenic focus assay for B cell cloning was used to analyze the responses of primary and secondary B cells obtained after fractionation on (1 × g) velocity sedimentation gradients. When nonimmune adult bone marrow cells are fractionated, the small cell (slowly sedimenting) fraction contains the majority of primary B cells specific for the 2,4-dinitrophenyl determinant (DNP). When nonimmune adult spleen cells are fractionated, all of the cell fractions contain B cells specific for DNP or for fluorescein (FL), and the large and medium cell fractions contain approximately 50 % of the activity. There is a differential expression of IgM vs. IgG1 anti-hapten monoclonal antibody, in that the majority of primary splenic B cells which give rise to clones producing only IgM antibody are found predominantly in the large and medium cell fractions. All of the cell fractions contain B cells which can generate clones producing both IgM and IgG1, or only IgG1 antibody. Ia-“negative” B cells which give rise to clones producing only IgM antibody are found in the large and medium cell fractions, whereas all the cell fractions contain la-positive B cells. When secondary spleen cells are fractionated, all of the cell fractions contain secondary B cell activity. The large and medium cell fractions contain half of the DNP-specific secondary B cells, whereas the medium and small cell fractions contain more secondary B cells specific for FL or hemocyanin. The large and medium cell fractions of secondary spleen cells are not enriched for B cells giving rise to clones producing only IgM antibody.