The in vitro immunogenicity of the solid-phase hapten, dinitrophenyl-ornithine-Bio-Gel (DNP-O-Bio-Gel), was investigated in cultures of mouse spleen cells. Appropriate combinations of cells and immobilized hapten were determined. Large numbers of direct anti-hapten plaque-forming cells (PFC) were generated when 1 × 107 C57BL/6 or C57BL/10 spleen cells were cultured with 4 × 103 DNP-O-Bio-Gel beads. Specificity studies of the responses of cultured spleen cells to DNP-O-Bio-Gel yielded the following results: soluble DNP-ornithine or DNP-bovine γ-globulin inhibited the induction of anti-hapten PFC by DNP-O-Bio-Gel; neither dinitrophenyl-Bio-Gel (DNP-Bio-Gel) nor ornithine-Bio-Gel (O-Bio-Gel) induced anti-hapten responsiveness; furthermore, neither DNP-Bio-Gel nor O-Bio-Gel inhibited the induction of PFC by DNP-O-Bio-Gel. It was concluded, from the results of these specificity experiments, that a spacer, ornithine, is required for immunogenicity of immobilized DNP; and that the Bio-Gel bead, itself, acts solely as a physical carrier for the hapten.

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