Regulation of Antibody Response in Vitro

Abstract

Abstract Rabbits were immunized by intraperitoneal injections of DNP-coupled ascaris antigen (DNP-Asc) precipitated with aluminum hydroxide gel, and the anamnestic antibody response of the mesenteric lymph node cells was studied in vitro. The cells were stimulated with homologous antigen, cultured in the absence of antigen for 6 days, and the IgM, IgG and IgE anti-DNP antibodies in the culture fluids were measured. In this system, formation of the anti-hapten antibodies was suppressed when the primed cells were stimulated with antigen in the presence of either ε-DNP lysine or ascaris antigen. At a given concentration of either the hapten or carrier, a similar extent of suppression was observed for the formation of the three immunoglobulins. The results suggest that both hapten-specific memory cells and carrier-specific helper cells are involved in the formation of anti-hapten antibodies of the three immunoglobulin classes. The primed lymph node cells gave maximal antibody response when they were stimulated with a certain concentration of DNP-Asc. The optimal concentration of the antigen for maximal response was found to depend upon the interval between the priming of donors and stimulation of primed cells. When the same cell suspension was employed, however, the optimal concentration of antigen was the same for the formation of IgG, IgM and IgE antibodies. The DNP-Asc-primed lymph node cells formed both IgM and IgG anti-hapten antibodies upon stimulation with DNP-bovine γG globulin (DNP-BGG), the amounts of the antibodies formed in the culture were only 10 to 20% of that formed by stimulation with DNP-Asc. The optimal concentration of the antigens for maximal antibody formation, and the ratio of IgM/IgG antibodies in the culture fluids was comparable whether DNP-Asc or DNP-BGG was used for stimulation. The results indicated that the optimal concentration of antigen and the distribution of anti-hapten antibodies between IgG and IgM classes is decided by the population of hapten-specific memory cells rather than carrier-specific helper cells. This idea was supported by the experiments in which rabbits primed with hapten primary-carrier conjugates received a supplemental immunization with a secondary carrier. The supplemental immunization enhanced the in vitro antibody response of their lymphocytes to the secondary carrier conjugates but did not influence the distribution of anti-hapten antibody between IgM and IgG. IgE antibody, however, was not formed in vitro when DNP-Asc-primed lymph node cells were stimulated by DNP-BGG, even when the donors had received supplemental immunization with BGG. The results indicated theat BGG-specific helper cells, which enhance both IgG and IgM antibody responses, did not collaborate with hapten-specific memory cells for IgE antibody formation.