Primary in vitro antibody response from human peripheral blood lymphocytes

Abstract

A method for the induction of a primary in vitro antibody response from human peripheral blood lymphocytes is presented. Upon cultivation with trinitrophenyl conjugated polyacrylamide beads (TNP-PAA), an anti-TNP response can be obtained as indicated by the appearance of direct plaque-forming cells from day 5 of culture, with a reproducible peak on day 8. These plaques correspond to cells actively producing antibody of the IgM type, as shown by their inhibition by cycloheximide and by anti-human IgM serum, but not by anti-human Fc gamma serum. Their specificity for the TNP hapten can be demonstrated by the effector cell blockade phenomenon, with highly substituted TNP-human IgG. Although the anti-TNP response induced by TNP-PAA in mouse spleen cell cultures appears T independent the same response in human PBL may involve in addition the participation of T cells, since E-RFC depletion before culture led to a markedly decreased number of plaque-forming cells. A significant response could be obtained from the PBL of all of the 30 normal individuals tested. Importantly, the response was reproducible in its magnitude in the six individuals tested in at least three different experiments. Thus, the in vitro stimulation of human PBL by TNP-PAA can be proposed as a reliable test for the study of human B cell function in a specific primary antibody response.