ntibody-mediated suppression of the immune response in vitro. 3. Low zone tolerance in vitro.

Immunological tolerance was induced by the exposure of mouse spleen cells to mixtures of polymerized flagellin and antibody in vitro for 6 hours, at 37°, prior to thorough washing and in vitro challenge with optimally immunogenic doses of polymerized flagellin. The induction of antibody-mediated tolerance depended critically on the ratio of antigen and antibody to which the cells were initially exposed. Low zone tolerance could be induced with subimmunogenic concentrations of monomeric flagellin, in amounts as low as 2 pg/ml (about 10-14 molar) and the appropriate concentration of antibody (a titre of 10-3). In contrast tolerance could not be induced with subimmunogenic concentrations of polymeric flagellin in the presence of a wide range of antibody doses. There was a remarkable similarity between the tolerant states obtained in vitro with tolerogenic doses of polymeric flagellin, and with immunogenic and subimmunogenic concentrations of flagellin in the presence of antibody, suggesting that the cellular basis of the tolerant state remains the same. An antigen-focussing hypothesis has been proposed to integrate the mechanisms of high zone and antibody-mediated tolerance at the cellular level.