A miniaturized cell-mediated cytotoxicity assay with human effector mononuclear cells

A miniaturized method (Microtest, MIT) for detecting natural killer (NK) and antigen-elicited cell-mediated cytotoxicity has been developed. It retains the sensitivity and the efficiency of conventional macroassay (Macrotest, MAT). In comparison with the standard MAT, MIT provides a 5-fold reduction in the number of effector and target cells without changing the final reaction volume. This avoids the excessive relative evaporation that could occur in microassays employing limited reaction volumes. Moreover the use of V-bottom microtiter plates allows the recovery of 0.15 ml of supernatant, thus increasing the efficiency of 51Cr recovery. MIT was adopted for the evaluation of the NK activity of untreated or interferon (IFN)-treated human mononuclear cells (MNC) and for cold-inhibition and cytotoxic T-lymphocyte (CTL) assays. In the experiments performed with both macro and micro assays, comparable values of the percentage of specific lysis and of the number of lytic units were found. The slopes of the curves obtained with MIT are generally slightly lower than those detectable with MAT. The Pearson coefficient r2 is generally better for the macroassay although it can be considered acceptable in the microassay. The MIT described here appears to be a useful method, especially for providing information on natural resistance and cytotoxic T-lymphocyte systems in a number of pathological conditions characterized by a small recovery of effector cells from standard blood collection for analytical purposes.