The measurement of natural killer (NK) cells toxicity against tumour or virus-infected cells especially in cases with small blood samples requires highly sensitive methods. Here, a coupled luminescent method (CLM) based on glyceraldehyde-3-phosphate dehydrogenase release from injured target cells was used to evaluate the cytotoxicity of interleukin-2activated NK cells against neuroblastoma cell lines. In contrast to most other methods, CLM does not require the pretreatment of target cells with labeling substances which could be toxicor radioactive. The effective killing of tumor cells was achieved by low effector/target ratios ranging from 0.5:1 to 4:1. CLM provides highly sensitive, safe, and fast procedure for measurement of NK cell activity with small blood samples such as those obtained from pediatric patients. (c) 2005 Elsevier Inc. All rights reserved.