THE USE OF NONRADIOACTIVE CHROMIUM AS AN ALTERNATIVE TO CR-51 IN NK ASSAY

A novel method to measure target cell cytolysis based on the use of 'cold', non-radioactive chromium and on the determination of metal release by graphite furnace atomic absorption spectroscopy (FAAS) is proposed. Natural killer (NK) assays were performed by labelling target cells with chromium as Na2CrO4, and results were compared with those obtained by conventional overnight labelling with Cr-51 of targets killed by the same effecters. The cytotoxic capacity of peripheral blood lymphocytes from healthy subjects was evaluated, and NK activity measured with both methods showed a good agreement at each of the tested effector to target cell ratios (between 100:1 and 1:1), with a high and significant coefficient of correlation (r = 0.931, p < 0.0001). The selection of the appropriate Cr concentrations for labelling target cells took into account both the sensitivity of our instrumentation and the possible toxic effects of the metal. A study of the effects of Cr on the cell line (K562) which is usually employed as a target in NK tests showed that Cr could have a detrimental effect on cellular function, with significant numbers of cells with depolarised mitochondria and reduced DNA synthesis after 24 h incubation using Cr levels higher than 15 mu mol/l (780 mu g/l). The method proposed here has a number of advantages, including the use of a non-radioactive tracer, limited costs, high sensitivity and reproducibility, and the possibility of storing samples. In addition, the technique uses a fixed Cr concentration which is known to be non toxic.