Saal J G, Rieber E P, Riethmüller G
Scand J Immunol. 1976;5(5):455-66. doi: 10.1111/j.1365-3083.1976.tb00300.x.
An in vitro lymphocyte-mediated cytotoxicity assay using [3H]proline-labeled target cells is described. The assay, modified from an original procedure of Bean et al., assesses the release of [3-H]proline by filtering the total culture fluid containing both trypsinized tumor cells and effector cells. Filtration is performed with a semiautomatic harvesting device using low suction pressure and large-diameter glass filters. Pretreatment of filters with whole serum diminishes adsorption of cell-free radioactive material considerably and thus increases the sensitivity of the assay. Nearly 100% of the radioactivity could be recovered with this harvesting device. The technique allowed the detection of cytolytic activities of lymphocytes after 6 h of incubation. Lymphocytes from patients with primary malignant melanoma showed a significantly higher cytolytic reactivity (P less than 0.001) than normal donors' lymphocytes against three different melanoma cell lines. In a series of parallel experiments on 36 patients and 18 normal donors, this modification of the [3H]proline test was compared with three different assays: the conventional microcytotoxicity test of Takasugi and Klein, the original [3H]proline microcytotoxicity test of Bean et al., and the viability count of tumor cells.
本文描述了一种使用[3H]脯氨酸标记靶细胞的体外淋巴细胞介导的细胞毒性试验。该试验是在Bean等人的原始方法基础上改进而来，通过过滤含有胰蛋白酶消化的肿瘤细胞和效应细胞的总培养液来评估[3H]脯氨酸的释放。过滤使用半自动收获装置，采用低吸力和大直径玻璃滤器进行。用全血清预处理滤器可显著减少无细胞放射性物质的吸附，从而提高试验的灵敏度。使用该收获装置可回收近100%的放射性。该技术能够在孵育6小时后检测淋巴细胞的细胞溶解活性。原发性恶性黑色素瘤患者的淋巴细胞对三种不同的黑色素瘤细胞系的细胞溶解反应性明显高于正常供体的淋巴细胞（P小于0.001）。在对36例患者和18名正常供体进行的一系列平行实验中，将这种[3H]脯氨酸试验的改进方法与三种不同的试验进行了比较：Takasugi和Klein的传统微细胞毒性试验、Bean等人的原始[3H]脯氨酸微细胞毒性试验以及肿瘤细胞的活力计数。
