NEW MONOCLONAL-ANTIBODIES DIRECTED AGAINST HUMAN RENIN - POWERFUL TOOLS FOR THE INVESTIGATION OF THE RENIN SYSTEM
NEW MONOCLONAL-ANTIBODIES DIRECTED AGAINST HUMAN RENIN - POWERFUL TOOLS FOR THE INVESTIGATION OF THE RENIN SYSTEM
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DOI：
10.1172/jci111488
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发表时间：
1984-01-01
影响因子：
15.9
通讯作者：
PAU, B
中科院分区：
文献类型：
--
作者：
GALEN, FX;DEVAUX, C;PAU, B
针对人肾素的单克隆抗体是通过将骨髓瘤细胞与注射了纯肿瘤或高度纯化肾素的 Balb/c 或高反应 Biozzi 小鼠的脾细胞融合而获得的。这些程序产生了 7 种针对人肾素的稳定单克隆抗体。通过在固相测定中与纯碘化肾素或不溶性肾素结合来筛选杂交瘤培养基中的抗体。提供与碘化肾素的50％结合的纯化抗体的浓度在1倍到1倍之间变化。 10-10 至 1 次。 10-7 M。发现两种单克隆抗体是体外肾素酶活性的有效抑制剂，表现为非竞争性抑制剂（Ki，1-4 × 10-10 M）。它们对灵长类肾素具有特异性。三种单克隆抗体为肾素纯化提供了合适的免疫吸附剂。其中一种免疫吸附剂用于大规模纯化肾酶，一步即可将肾素富集 825 倍。两种抗体能够区分活性和非活性肾素，并能够同时分离和纯化各种生物液体中的两种酶形式。当使用间接免疫荧光和过氧化物酶-抗过氧化物酶技术时，单克隆抗体也对人和猴肾素进行染色。用 2 种单克隆抗体构建了高灵敏度的肾素放射免疫测定法。这种改进的测定法的灵敏度应该允许检测正常人血浆中的肾素。单克隆抗体已被证明在以下领域优于多克隆抗体：活性肾素与非活性肾素的分离、从生物体液中纯化肾素以及建立血浆肾素的直接测定。
Monoclonal antibodies directed against human renin were obtained by the fusing of myeloma cells with spleen cells from Balb/c or high-responder Biozzi mice injected with pure tumoral or highly purified renal renin. These procedures resulted in the production of 7 stable monoclonal antibodies to human renin. Antibodies in the hybridoma culture medium were screened by binding to pure iodinated renin or insolubilized renin in a solid phase assay. The concentration of purified antibodies that provided a 50% binding to iodinated renin varied from 1 .times. 10-10 to 1 .times. 10-7 M. Two monoclonal antibodies were found to be potent inhibitors of renin enzymatic activity in vitro, behaving as noncompetitive inhibitors (Ki, 1-4 .times. 10-10 M). They were specific for primate renin. Three monoclonal antibodies provided suitable immunoadsorbants for renin purification. One of these immunoadsorbants was used for large-scale purification of the renal enzyme, resulting in an 825-fold renin enrichment in a single step. Two antibodies were able to distinguish between active and inactive renin, and enabled concomitant separation and purification of the 2 enzyme forms in various biological fluids. Monoclonal antibodies also stained human and monkey renal renin when indirect immunofluorescence and peroxidase-antiperoxidase techniques were used. A highly sensitive radioimmunometric assay of renin was constructed with 2 monoclonal antibodies. The sensitivity of this improved assay should permit the detection of renin in normal human plasma. Monoclonal antibodies have been shown to be superior to polyclonal antibodies in the following areas: the separation of active from inactive renin, the purification of renin from biological fluids and the setting up of a direct assay of plasma renin.