RAPID COLORMETRIC ASSAY FOR CELL VIABILITY - APPLICATION TO THE QUANTITATION OF CYTO-TOXIC AND GROWTH INHIBITORY LYMPHOKINES
RAPID COLORMETRIC ASSAY FOR CELL VIABILITY - APPLICATION TO THE QUANTITATION OF CYTO-TOXIC AND GROWTH INHIBITORY LYMPHOKINES
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DOI：
10.1016/0022-1759(84)90190-x
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发表时间：
1984-01-01
影响因子：
2.2
通讯作者：
WARE, CF
中科院分区：
文献类型：
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作者：
GREEN, LM;READE, JL;WARE, CF
建立了一种快速比色微量滴定法，用于检测凝集素或肿瘤细胞激活的人淋巴细胞产生的细胞毒性淋巴因子。使用四唑染料检测经光敏素处理的靶细胞的活力，所述四唑染料被活细胞而非死细胞还原成蓝色甲瓒。使用微孔板分光光度计（ELISA [酶联免疫吸附试验]酶标仪）定量形成的染料量，目视观察证实产生的甲瓒染料量与活靶细胞数量成正比。使用这种比色法的优点是它不需要洗涤步骤或放射性同位素，并且具有精确性和快速性。使用小鼠L929 [肿瘤成纤维细胞]和人ESH-5L细胞系作为检测人淋巴细胞产生的光毒素的靶细胞，建立了最佳条件。L929细胞系比ESH-5L细胞系对由人植物凝集素-P激活的T淋巴细胞产生的细胞毒素或由各种肿瘤细胞系刺激的外周血淋巴细胞产生的细胞毒素的裂解活性敏感10至50倍。该测定系统也可用于检测能够中和光毒素活性的抗体，并且应该是有助于这些淋巴因子的分子表征的合适方法。
A rapid colorimetric microtiter assay was developed to detect cytotoxic lymphokines produced by human lymphocytes activated with lectins or tumor cells. The viability of lymphotoxin-treated target cells was detected using a tetrazolium dye that is reduced to a blue formazan by living but not dead cells. The amount of dye formed was quantitated using a microplate spectrophotometer (ELISA [enzyme-linked immunosorbent assay] plate reader) and visual observations confirmed the amount of formazan dye produced was directly proportional to the number of viable target cells. The advantages of using this colormetric method are that it requires no washing steps or radioisotopes and its precision and rapidity. Optimal conditions were established using the murine L929 [neoplastic fibroblasts] and human ESH-5L cell lines as target cells for detecting lymphotoxins produced by human lymphocytes. The L929 cell line was 10- to 50-fold more sensitive than the ESH-5L line to the lytic activity of cytotoxins produced by human phytohemagglutinin-P-activated T lymphocytes, or the cytotoxins produced by peripheral blood lymphocytes stimulated with various tumor cell lines. This assay system was also useful in detecting antibodies capable of neutralizing lymphotoxin activity and should be a suitable method to aid in the molecular characterization of these lymphokines.