Detection of antimitochondrial autoantibodies in immunofluorescent AMA-negative patients with primary biliary cirrhosis using recombinant autoantigens
Detection of antimitochondrial autoantibodies in immunofluorescent AMA-negative patients with primary biliary cirrhosis using recombinant autoantigens
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DOI：
10.1053/jhep.2001.26514
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发表时间：
2001-08-01
期刊：
影响因子：
13.5
通讯作者：
Gershwin, ME
中科院分区：
文献类型：
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作者：
Miyakawa, H;Tanaka, A;Gershwin, ME
抗线粒体抗体（AMA）是原发性胆汁性肝硬化（PBC）的血清学标志。然而，根据临床实验室的不同，使用天然线粒体抗原和包括免疫荧光 (IMF) 和酶联免疫吸附测定 (ELISA) 在内的各种常规检测，5% 至 17% 的 PBC 患者始终呈 AMA 阴性。主要的免疫反应性线粒体自身抗原是2-含氧酸脱氢酶复合物家族的E2成员，包括丙酮酸脱氢酶复合物-E2 (PDC-E2)、支链2-含氧酸脱氢酶复合物-E2 (BCOADC-E2)和含氧戊二酸脱氢酶复合物-E2 (OGDC-E2)；这些蛋白质的 cDNA 现已被克隆、测序，并确定了它们的 B 细胞表位。在本研究中，我们从人类而非牛中克隆了编码这些蛋白质的 cDNA，并在新开发的 ELISA 中使用独特的大肠杆菌缓冲液表达了重组蛋白，并将数据与之前使用 AMA 阳性和阴性患者进行的检测进行了比较。使用这种新的检测方法以及我们的阳性标准（光密度 (OD) 高于对照血清平均值 10 SD），191 份 PBC 血清的 AMA 阳性率为 94%（191 份中的 179 份），而 IMF 的阳性率为 84%（191 份中的 161 份）。 316 份对照血清均无反应。使用我们的重组检测，我们将注意力集中在 30 名 IMF-AMA 阴性患者上。使用这种新的 ELISA，30 名患者中有 22 名 (73%) 呈阳性。由 30 名 IMF-AMA 阴性/ ELISA 阳性患者组成的组在年龄、性别分布、肝功能测试、血清 IgM 升高或病理分期方面与 IMF-AMA 阳性患者的可比较群体没有显着差异。
Antimitochondrial antibodies (AMA) are the serologic hallmark of primary biliary cirrhosis (PBC). However, depending on the clinical laboratory, from 5% to 17% of PBC patients are consistently AMA-negative, using native mitochondrial antigens and a variety of conventional assays including immunofluorescence (IMF) and enzyme-linked immunosorbent assay (ELISA). The major immunoreactive mitochondrial autoantigens are the E2 members of the 2-oxo-acid dehydrogenase complex family, including pyruvate dehydrogenase complex-E2 (PDC-E2), branched chain 2-oxo acid dehydrogenase complex-E2 (BCOADC-E2), and oxo-glutarate dehydrogenase complex-E2 (OGDC-E2); cDNAs of these proteins have now been cloned, sequenced, and their B-cell epitopes defined. In the present study, we cloned cDNAs encoding these proteins from human, not bovine, sources, and expressed the recombinant proteins in a newly developed ELISA that employs a unique Escherichia coli buffer, and compared the data with previous assays using both AMA-positive and -negative patients. Using this new assay and our criteria for positive as an optical density (OD) greater than 10 SD above the mean of control sera, the AMA-positive rate of 191 PBC sera was 94% (179 of 191) compared with 84% (161 of 191) by IMF. None of the 316 control sera were reactive. Using our recombinant assays, we focused attention on the 30 IMF-AMA-negative patients. Twenty-two of 30 (73%) of these patients were positive using this new ELISA. The group of 30 IMF-AMA-negative/ ELISA-positive patients did not differ significantly from a comparable population of IMF-AMA-positive patients with respect to age, sex distribution, liver function tests, elevation of serum IgM, or pathologic stage.