Quantitative measurement of T-lymphocyte activation by an enzyme-linked immunosorbent assay (ELISA) detecting interleukin-2 receptor expression

Abstract

A monoclonal antibody prepared against the murine interleukin-2 receptor (IL-2R) was employed to develop an ELISA method for measuring the immunological activation of T-cells. The assay detects an increase in IL-2R expression on activated lymphocytes. Stimulated splenic lymphocytes displayed markedly higher IL-2R expression compared to unstimulated controls. A significant increase in IL-2R expression on lymphocytes was detected in mitogen-stimulated responses, in a one-way mixed leukocyte reaction (MLR) and in the antigen-specific responses to conalbumin and purified protein derivative (PPD) in vitro. At a constant cell number, the level of IL-2R expression was found to be dependent on the dose of the stimulant. A comparative study of the kinetics of activation of splenic lypphocytes in response to mitogen, antigen and allogeneic cells as measured by the IL-2R ELISA and the conventional tritiated thymidine ( 3HTdR) uptake assay revealed remarkable similarity. For both assays, the mitogenic response was detected within 12 h ,the peaked at 72 h, and MLR was detectable within 2–3 days and peaked at day 6, and the specific antigenic response was detected within 2 days and peaked on day 4–5. Hydroxyurea, an inhibitor of DNA synthesis, had no effect on early IL-2R expression by mitogen-stimulated splenic lymphocytes, however, only 20% of maximum IL-2R expression could be detected at later stages of incubation. In contrast, cycloheximide, an inhibitor of protein synthesis, completely abrogated IL-2R expression and proliferation of stimulated lymphocytes.