细胞毒效应的DNA荧光测定法测定恶性肿瘤患者外周血NK细胞活性

DNA-fluorometric assay for cytotoxicity and NK cytotoxic activety of peripheral blood lymp?o cytes from cancer patients

本文根据溴化乙锭(EB)可特异地与双链DNA嵌合产生荧光反应的特性,建立了细胞毒效应的DNA荧光测定法。本法的主要特点是通过测定效应细胞和靶细胞作用后残留靶细胞DNA的荧光强度和对照靶细胞的DNA荧光强度,分析效应细胞对靶细胞的杀伤效应。研究结果表明,人外周血淋巴细胞与人红白血病K562靶细胞按不同的效靶比例共同培养20小时后,未损伤的残留靶细胞DNA的荧光强度和对照靶细胞DNA的荧光强度的比值与效应细胞的杀伤效应呈负相关性。本文检测了34例正常人和30例恶性肿瘤患者外周血淋巴细胞的NK细胞活性,其平均值分别为51.96±6.62%和25.02±10.83%两者之间有非常显著的差异(P<0.01),表明用DNA荧光测定法检测NK细胞对K562靶细胞的杀伤活性具有较高的敏感性。

On the fluorescent reaction of ethidium bromide binding double-stranded DNA after cell ly-sis has been used to develop a rapid fluorometric assay for cytotoxicity. By using this technique,the cytotoxic activity of effector cells against target cells can be assayed by measuring the fluo-rescent intensities of the residual target cell DNA and the control target cell DNA. Our experi-mental results show that the fluorescent intensity of the uninjured, residual K562 cell DNA is ininverse relation to the cytotoxic activity of NK cells after incubation of peripheral bloed lympho-cytes with K562 cells. for 20 hr, at a different ratio of effector cell to target cell. We detected thecytotoxic activities of NK cells of perpheral blood lymphocytes from 34 normal individuals and 30cancer patients, and their mean percentages of NK cytotoxicity were 51.96±6.62% and 25.02±10.83% respectively, and had much significant deviation statistically(P<0.01). These resultsindicate that the DNA-fluorometric assay for NK cytotoxic activity has a high degree of precisionand sensitivity, and an excellent reproducibility (CV=4.8%). The main advantages of this assayare its simplicity, rapidility, and the lack of any radioisotope for detection of cytotoxicity.