NONRADIOACTIVE ALTERNATIVE TO CLINICAL MIXED LYMPHOCYTE-REACTION

The MLR is used clinically as a functional assay for genetic HLA dispa rity. Traditionally this test relies on [H-3]thymidine incorporation t o detect T-cell proliferation as an indicator of alloantigenicity. Env ironmental and administrative concerns regarding radioisotope use and disposal have encouraged the development of sensitive, nonradioactive assay for T-cell alloactivation. We describe a nonradioactive alternat ive to the clinical MLR which uses an IL-2-dependent cell line, CTLL20 .3, and MTT reduction to detect IL-2 accumulation in MLC SNs as an ind ex of T-cell alloactivation. We first determined (a) the optimal numbe r of CTLL20.3 cells for the assay, (b) the optimal time of SN analysis for IL-2, and (c) additional manipulations that significantly increas e the assay sensitivity for IL-2. Using this assay system with patient lymphocyte combinations, we demonstrated that the CTLL20.3-MTT assays correlate well with the [H-3]thymidine assays of T-cell proliferation for the detection of MHC incompatibility. Indeed, the CTLL20.3-MTT as say may be slightly more sensitive than the traditional clinical MLR.