A Cytolytic System for the in Vitro Detection of Cell-Mediated Immunity to Soluble Antigen

Abstract

An in vitro cytotoxic test is described for the demonstration of cell-mediated immunity to soluble antigen. This system has been used to study the development of a cytotoxic splenic lymphocyte population in the guinea pig after immunization with dinitrophenylated human IgG (DNP-HGG). As “target” cells, ⁵¹Cr-labeled mouse mastocytoma cells were used, and were coupled to DNP-HGG with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide HCl. “Target” cell destruction was assessed by comparing the rates of ⁵¹Cr release from these cells in the presence of splenic lymphocytes from specifically sensitized animals with that released in the presence of the same number of lymphocytes from unimmunized animals. The mastocytoma cells were found to be preferable to erythrocytes as carriers of antigen, in that they were more sensitive to membrane permeability changes. Splenic lymphocytes from guinea pigs immunized with DNP-HGG caused specific lysis of DNP-HGG-coated mastocytoma cells. Cells coated with DNP-BSA and HGG were lysed to a lesser extent, while cells coated with antigens immunochemically unrelated to the antigen used for sensitization were not lysed to any greater extent than that observed with lymphocytes from unimmunized animals. Several factors distinguished cytolysis by “sensitized” lymphocytes from that mediated by serum antibody in the presence of complement. Cytolysis by lymphocytes was not inhibited by anti-guinea pig immunoglobulin antisera. The total cytolytic activity of splenic lymphocytes did not parallel measured serum antibody. The specificity of cytolysis by splenic lymphocytes from animals immunized with DNP-HGG was markedly carrier-dependent.