Polyclonal B cell responses in the presence of defined filler cells: complementary effects of lipopolysaccharide and anti-immunoglobulin antibodies.

The signal requirement for polyclonal B cell responses in the presence of T helper (Th) cells, lipopolysaccharide (LPS), anti-Ig antibodies coupled to Sepharose beads (anti-Ig) and/or T cell supernatants (SN) was studied in a murine system using (a) low numbers of B cells/culture in order to reduce the effects of contaminating Th cells and (b) defined sources of irradiated filler cells in the form of EL4 thymoma cells or cloned male H-Y antigen-specific Th cells. The results demonstrate that for optimal proliferation as well as (protein A) plaque-forming cell (PFC) generation B cells required at least two activation signals in addition to factor(s) present in Th or EL4 SN, i.e. either a specific (or concanavalin A-dependent nonspecific) Th signal and anti-Ig or (in cultures with EL4 filler cells) LPS and anti-Ig. While confirming several previous studies, including our own, which showed a requirement for two activation signals in conjunction with T cell factors in antigen-specific B cell responses, the present system differs from previous polyclonal systems by showing nonoverlapping effects of LPS or a specific Th signal on the one hand and anti-Ig on the other in the induction of growth factor responsiveness of B cells. In addition, limiting dilution analysis showed that in cultures with EL4 filler cells, LPS, anti-Ig and EL4 SN 1/8 surface Ig-positive cells generated greater than 10 PFC with a mean clone size of 70 PFC and indicated that only the B cells were limiting. This system using defined thymoma filler cells should be useful for assaying factors potentially replacing the LPS or anti-Ig signals.