De Groote D, Marchant A, Fauchet F, Jadoul M, Dehart I, Gérard C, Gevaert Y, Lopez M, Gathy R, Franssen J D
Department of Endocrinology, University of Liège, Belgium.
J Immunol Methods. 1994 Dec 28;177(1-2):225-34. doi: 10.1016/0022-1759(94)90160-0.
Biological and biochemical characteristics of monoclonal antibodies (MABs) raised against human interleukin-10 (IL-10) are described as well as their use in the design of a specific ELISA for the measurement of the cytokine. 21 murine anti-human interleukin-10 (IL-10) MABs were obtained by fusion of splenocytes from mice immunized against human recombinant IL-10 with SP2/0 myelomatous cells. These antibodies define three major antigenic areas on the IL-10 molecule, one of which comprises epitopes involved in receptor binding and induction of biological activity. They recognize recombinant human IL-10 with affinities ranging from 1.3 x 10(-7) to 3 x 10(-11), as well as natural IL-10. Most of them also recognize viral IL-10 (vIL-10) encoded by the Epstein-Barr virus (EBV). A specific human-IL-10 ELISA has been developed using two MABs (18 and 19) as capture antibody and one MAB (17) as detector. The sensitivity (3 pg/ml), precision (intra-assays < 4%), reproducibility (interassay < 3%), and accuracy (recoveries, ranging between 84 and 107%, in several fluids) of the assay, plus its excellent performance in dilution tests, and the lack of interference when in the presence of possible cross-reactive substances, permits accurate cytokine measurement in biological fluids such as serum, plasma, bronchoalveolar lavage, urine and culture supernatants. Using the assay, IL-10 was measurable in the plasma of patients with septic shock (range 11-2740 pg/ml) whereas IL-10 plasma levels were < 7.8 pg/ml in healthy volunteers.
本文描述了针对人白细胞介素 -10（IL -10）产生的单克隆抗体（MABs）的生物学和生化特性，以及它们在设计用于检测该细胞因子的特异性酶联免疫吸附测定（ELISA）中的应用。通过将免疫了人重组IL -10的小鼠脾细胞与SP2/0骨髓瘤细胞融合，获得了21种鼠抗人白细胞介素 -10（IL -10）单克隆抗体。这些抗体确定了IL -10分子上的三个主要抗原区域，其中一个区域包含参与受体结合和生物活性诱导的表位。它们识别重组人IL -10的亲和力范围为1.3×10⁻⁷至3×10⁻¹¹，也识别天然IL -10。它们中的大多数还识别由爱泼斯坦 - 巴尔病毒（EBV）编码的病毒IL -10（vIL -10）。已经开发出一种特异性人IL -10 ELISA，使用两种单克隆抗体（18和19）作为捕获抗体，一种单克隆抗体（17）作为检测抗体。该测定的灵敏度（3 pg/ml）、精密度（批内<4%）、重现性（批间<3%）和准确性（在几种液体中的回收率在84%至107%之间），加上其在稀释试验中的优异性能，以及在可能存在交叉反应物质时缺乏干扰，使得能够在血清、血浆、支气管肺泡灌洗、尿液和培养上清液等生物液体中准确测量细胞因子。使用该测定法，在感染性休克患者的血浆中可检测到IL -10（范围为11 - 2740 pg/ml），而健康志愿者的血浆IL -10水平<7.8 pg/ml。
