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Rosette plaques with lymphoid cells from heterozygous rabbits. Ani-hapten antibody-forming cells displaying either
Abstract:
A rosette-plaque model was employed to test for the expression of b locus allelic markers at the surface of lymph node lymphocytes (LNL) from heterozygous (b4,6) rabbits, 5 days after immunization with NIP-diphtheria toxoid. Before immunization, in all animals examined, LNL displaying both b4 and b6 determinants at the surface (range 5-20 per cent) were detected, the remainder consisting of cells exhibiting only one or the other determinant. After immunization, five of the thirteen heterozygotes apparently had gone into allelic exclusion as LNL from these animals showed only b4 or b6 rosettes which secreted anti-NIP antibody in the plaque. The eight remaining rabbits remained in allelic inclusion. Since cytophilic uptake of allotype might have contributed to double expression, LNL from immunized animals were treated with pronase to remove surface immunoglobulin. When the stripped cells were cultured overnight in serum-free medium, reappearance of b4, b6, and b4 plus b6 expressing cells was seen. When pronase-stripped cells were incubated in cycloheximide (20 mug/ml) for 5 hr, no allotype synthesis was found but inhibition was relieved when the cells were washed free of the antibiotic. Regrowth resulted in rosette levels similar to those found originally in the three allotype-bearing populations. Stripping the cell surface allotype with pronase, and allowing regrowth of allotype overnight also resulted in one of four animals regaining the ability to express both allotypes at the surface in the plaque-forming situation. Lymphocytes from homozygous controls (b4,4 and b6,6) displayed their own individual allelic markers either when the cells from each were tested alone or in combination, unimmunized or immunized. An additional finding was the apparent lack of allelic preference for NIP in the heterozygotes as approximately similar numbers of cells were found bearing the b4 and b6 marker at the surface in the NIP plaque.