Requirement for persistent extracellular antigen in cultures of antigen-binding B lymphocytes

A system was established to assess the requirement for continuous presence of antigen in B-lymphocyte activation to antibody formation. Mouse spleen B lymphocytes, enriched for cells bearing anti-NIP (hapten 4-hydroxy-3-iodo-5-nitrophenylacetic acid) receptors, were pretreated briefly with NIP-POL (polymerized flagellin) antigen, washed, and added in small numbers to microcultures. The behaviour of these cells was compared with that of cells cultured in the continuous presence of antigen. Unfractionated spleen cells were studied under similar conditions. In contrast to unfractionated cells, enriched cells could not be triggered effectively by brief contact with antigen at any concentration tested. Fewer cells were activated, and clone size was smaller after brief contact with antigen than when antigen was present continuously. Furthermore, brief contact at high concentration did not cause tolerance induction.