An analysis of variables affecting the measurement of tumor immunity in vitro with 125I-iododeoxyuridine-labelled target cells. Studies of immunity to primary Moloney sarcomas.

Abstract

AbstractVariables affecting the measurement of immunity to adherent tumor cells in vitro have been analyzed using the immune response to primary Moloney sarcomas in BALB/c mice as a model system. IUdR, a gamma‐emitting isotope which is incorporated into the DNA of dividing tumor cells, was used to quantitate the number of adherent tumor cells in 0.2 ml cultures. Lymphoid cell‐mediated killing of tumor cells was measured by labelling tumor cells before they were cultured with effector cells (cytotoxicity test), and lymphoid cell‐mediated inhibition of tumor‐cell growth was measured by labelling those tumor cells surviving interactions with effector cells (cytostasis test). Addition of lymphoid cells to adherent tumor cells initiates complex interactions which have non‐specific (i.e., unrelated to recognition of tumor antigens) and specific (i.e., related to recognition of tumor antigens) effects on the survival and growth of tumor cells. The magnitude of these effects, as measured in the cytotoxicity or cytostasis tests, depends upon the number, kind, and state of activation and/or sensitization of effector cells and upon the number and susceptibility of tumor cells. Effector cells from control mice have only non‐specific effects whereas those from MSV‐immunized mice have both non‐specific and specific effects. Lymph‐node cells (LNC) from normal control mice non‐specifically kill IUdR labelled tumor cells in the cytotoxicity test, whereas killing and growth‐promoting influences are both present in the cytostasis test. Non‐specific cytotoxic effects also were mediated by LNC from MSV‐immunized mice, and these effects were of greater magnitude than those mediated by LNC from normal control mice. Our present and previous experiments suggest that non‐specific cytotoxicity of LNC is largely due to macrophages. In contrast, non‐specific promotion of tumor‐cell growth in the cytostasis test is apparently mediated by lymphocytes. Specific cytotoxicity or cytostasis was obtained only when non‐adherent LNC (NALNC) containing 99.8% lymphocytes and less than 0.2% macrophages were employed as effector cells. Although both cytotoxicity and cytostasis tests seem to give similar results, we conclude, for a number of reasons, that the cytotoxicity test is preferable for measuring cell‐mediated immune responses to adherent tumor cells. Our findings and conclusions should be generally applicable to studies of animal and human tumors.