Absence of Agglutinin Formation in Spleen Cell Cultures from Immunologically Tolerant Mice

Summary

1. 1. Mice injected with a tolerance-inducing concentration of Shigella soluble antigen at birth formed only low or negligible levels of detectable serum agglutinins upon subsequent immunization with Shigella antigen at 5 to 6 weeks of age. Control mice not treated at birth formed readily detectable levels of agglutinins upon similar challenge immunization.

2. 2. Spleen cell suspensions from Shigella tolerant mice incubated in tissue culture medium for a period of 1 to 2 weeks failed to secrete detectable amounts of Shigella agglutinins after either in vivo or in vitro stimulation with antigen. Spleen cell suspensions from normal mice injected with Shigella antigen 5 to 10 days prior to sacrifice formed readily measurable levels of agglutinins in vitro under the same conditions during the same time period.

3. 3. The inability of spleen cell cultures from Shigella tolerant mice to form agglutinins in vitro was antigen specific, since spleen cell cultures from either Shigella tolerant or normal control mice were capable of forming hemagglutinins to sheep erythrocytes in vitro following in vivo challenge immunization with sheep RBC′s.

4. 4. Hemagglutinin formation to sheep erythrocytes by cell suspensions from both tolerant and normal mice could be enhanced by in vitro stimulation with S-RBC. Hemagglutinin responses were also enhanced in cultures from both tolerant and normal animals by in vivo injection with both S-RBC and Shigella antigen, suggesting an adjuvant effect for endotoxin-containing Shigella antigen even in tolerant mice.

5. 5. The absence of agglutinins in spleen cell culture supernatants from tolerant mice was apparently due to central failure of specific agglutinin formation, since spleen cells from tolerant animals did not interfere with agglutinin formation when added to normal spleen cell cultures.

6. 6. Spleen cell cultures prepared from 16- to 40-week-old mice injected at birth with Shigella were capable of forming detectable levels of agglutinins in vitro, suggesting that the animals had “escaped” from the tolerant state.