HUMORAL IMMUNE-RESPONSE TO SALMONELLA-ABORTUSOVIS IN SHEEP - IN-VITROINDUCTION OF AN ANTIBODY-SYNTHESIS FROM EITHER SENSITIZED OR UNPRIMEDLYMPH-NODE CELLS

In vitro culture conditions were determined to induce an anti-Salmonel la abortusovis antibody synthesis from lymph node leucocytes of three immunized sheep and two unprimed lambs maintained in culture in the pr esence of heat-inactivated bacteria for 2 weeks. Humoral immune respon ses were assessed by enumerating specific antibody-secreting cells usi ng ELISASPOT and by titrating immunoglobulins secreted into culture su pernatants using ELISA techniques. Optimal secondary antibody response was observed from cultures performed with fetal calf serum (compared with horse serum) and with an antigen concentration of one to ten bact eria per cell. This kind of antigenic stimulation allowed induction of numerous antibody-secreting cells without adsorption of the secreted antibodies. Maximal numbers of antibody-secreting cells could reach a rate of 1% of the sheep leucocytes initially put into culture. Kinetic profiles of antibody production from boosted lymph node cells were ch aracterized by an ascending phase from the sixth to the twelfth day of culture and then showed a plateau phase until Day 14. Most of the res ponses were composed of IgM and IgG1 antibodies, traces of IgG2 being detected at the end of experiments. From the twelfth day of antigenic stimulation, the IgM isotype was preferentially expressed with high an tigen concentration (100 bacteria per cell), whereas the highest amoun ts of IgG1 were detected at lower concentration (one to ten bacteria p er cell). While anti-Salmonella IgM appeared to be mainly specific for the lipopolysaccharide (LPS) cell wall fraction, some IgG1 recognized other bacterial antigens. Kinetic profiles and magnitudes of primary antibody responses induced in vitro from lamb lymph node cells did not differ from those defined in cultures of sheep boosted leucocytes. Bu t these immune reactions were mainly made up of anti-LPS IgM. Few anti -Salmonella IgG1 were detected from the tenth day of culture. So these in vitro assays allowed induction of an antibody synthesis from eithe r in vivo sensitized or unprimed sheep lymph node leucocytes. This met hodology would permit achievement of more detailed studies on interact ions between Salmonella and lymph node leucocytes, leading to a better understanding of the mechanisms controlling bacterial dissemination t hrough the lymphoid tissue.