Production and characterization of monoclonal antibodies to human interleukin 2: strategy and tactics.
Production and characterization of monoclonal antibodies to human interleukin 2: strategy and tactics.
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DOI：
10.4049/jimmunol.131.4.1808
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发表时间：
1983-10
影响因子：
4.4
通讯作者：
K. Smith;M. Favata;S. Oroszlan
中科院分区：
文献类型：
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作者：
K. Smith;M. Favata;S. Oroszlan
人类白细胞介素2 （IL - 2）独特的激素特征，主要是IL - 2受体相互作用的高亲和力，对这种淋巴因子的单克隆抗体的产生产生了几个障碍。由于正常细胞来源每升只能产生几微克的IL - 2，因此有必要利用人T白血病细胞系的高生产者克隆和亚克隆来获得免疫原性的IL - 2蛋白。此外，需要抗体介导干预高亲和力IL - 2受体结合的检测方法在鉴定抗IL - 2产生杂交瘤方面是无效的，因此需要开发免疫检测方法。酶联免疫分析法检测到的三种最初衍生的抗体中有两种被发现与IL - 2特异性反应，这是由抗体浓度依赖的IL - 2活性中和所证明的。细胞增殖的中和作用对IL - 2反应细胞是特异性的，与抑制IL - 2受体结合相一致，可以被亲和纯化的IL - 2完全克服，并且是种特异性的；人和小鼠IL - 2被中和，而大鼠IL - 2活性不受影响。第三种抗体虽然在中和IL - 2活性方面效果较差，但能更有效地与IL - 2结合，并作为有效的免疫吸收剂发挥作用。利用免疫吸附法可一步富集纯化IL - 2。经十二烷基硫酸钠聚丙烯酰胺凝胶电泳、反相液相色谱和氨基末端氨基酸序列分析，纯化后的产物由单个蛋白质组成（Mr = 15,500），保留了生物活性。这些结果表明，个体淋巴因子的独特生化和功能特性可能很好地决定了抗体被激发和检测的效率。然而，抗淋巴因子一旦产生，就非常适合于探索这些免疫调节分子的分子和生物学特性。
The unique hormonal characteristics of human interleukin 2 (IL 2), primarily the high affinity of the IL 2-receptor interaction, created several impediments to the generation of monoclonal antibodies to this lymphokine. Because normal cell sources produce only a few micrograms of IL 2 per liter, it was necessary to utilize high producer clones and subclones of a human T leukemia cell line to obtain immunogenic amounts of IL 2 protein. Moreover, assays that required antibody-mediated intervention of the high affinity IL 2-receptor binding were ineffectual in the identification of anti-IL 2-producing hybridomas, thus necessitating the development of immunoassays. Two of three initially derived antibodies detected by enzyme-linked immunoassay were found to react specifically with IL 2 as demonstrated by antibody concentration-dependent neutralization of IL 2 activity. The neutralization of cellular proliferation was specific for IL 2-reactive cells, coincided with an inhibition of IL 2 receptor binding, could be completely overcome by affinity-purified IL 2 and was species-specific; human and murine IL 2 were neutralized, whereas rat IL 2 activity remained unaffected. A third antibody, although much less effective in neutralizing IL 2 activity, bound to IL 2 more avidly and functioned as an efficient immunoabsorbent. IL 2 could be concentrated and purified by immunoabsorption from crude conditioned medium in a single step. The purified product, which retained biologic activity, was made up of a single protein (Mr = 15,500) as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis, reversed-phase liquid chromatography, and amino-terminal amino acid sequence analysis. These results indicate that the distinctive biochemical and functional properties of individual lymphokines may well determine the efficiency with which antibodies may be elicited and detected. However, once produced, anti-lymphokines are uniquely suited for the exploration of the molecular and biologic properties of these immunoregulatory molecules.