Change of cytokeratin organization during development of Mallory bodies as revealed by a monoclonal antibody.
Change of cytokeratin organization during development of Mallory bodies as revealed by a monoclonal antibody.
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单克隆抗体揭示了马洛里体发育过程中细胞角蛋白组织的变化。
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发表时间：
1986
期刊：
影响因子：
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通讯作者：
D. Schiller
中科院分区：
文献类型：
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作者：
R. Hazan;Hüseyin Denk;W. Franke;E. Lackinger;D. Schiller
从灰黄霉素处理的小鼠的肝组织中分离出马洛里体（MB）物质，制备了鼠单克隆抗体（KM 54-5）。通过MB的阳性免疫荧光显微镜和通过十二烷基硫酸钠聚丙烯酰胺凝胶电泳分离的多肽上的免疫印迹技术鉴定抗原。在免疫印迹实验中，抗体KM 54-5与鼠、牛和人肝细胞的细胞角蛋白A（人8号）和D（人18号）以及分离的鼠MB中存在的细胞角蛋白A（8号）及其降解产物反应。在免疫荧光显微镜下，该抗体不与正常肝脏的细胞角蛋白丝反应，但在MB发育达到一定阶段后与MB呈阳性反应。在斑点印迹试验中，使用从鼠肝脏分离并在含有8 M尿素的pH 8缓冲液中通过离子交换色谱法纯化的单个细胞角蛋白多肽，抗体与单个多肽A（编号8）和D（编号18）反应，但不与在4 M尿素中从这些多肽重构的异型四聚体（A2 D2）反应。这些发现证实了MB细丝的细胞角蛋白性质。此外，他们表明MB形成的病理过程涉及细胞角蛋白组织和构象的变化，导致在异型四聚体亚基和正常细胞的细胞角蛋白丝中无法接近（“掩蔽”）的特定抗原决定簇的可接近性。因此，这项研究提出了一个例子的病理变化的细胞角蛋白丝，并说明了价值的单克隆抗体在检测这种变化。
A monoclonal murine antibody (KM 54-5) was produced against Mallory body (MB) material isolated from liver tissue of griseofulvin treated mice. The antigen was identified by positive immunofluroescence microscopy of MBs and by the immunoblotting technique on polypeptides separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis. In immunoblotting experiments, antibody KM 54-5 reacted with cytokeratins A (human no. 8) and D (human no. 18) of murine, bovine, and human hepatocytes as well as with cytokeratin A (no. 8) and its degradation products present in isolated murine MB. In immunofluorescence microscopy the antibody did not react with cytokeratin filaments of normal liver but showed a positive reaction with MBs after a certain stage in MB development had been reached. In a dot blot assay, using individual cytokeratin polypeptides isolated from murine liver and purified by ion exchange chromatography in pH 8 buffer containing 8 M urea, the antibody reacted with the individual polypeptides A (no. 8) and D (no. 18) but not with the heterotypic tetramer (A2D2) reconstituted from these polypeptides in 4 M urea. These findings confirm the cytokeratin nature of MB filaments. In addition, they show that the pathologic process of MB formation involves changes in cytokeratin organization and conformation, resulting in the accessibility of a specific antigenic determinant which is inaccessible ("masked") in the heterotypic tetramer subunit and in the cytokeratin filaments of normal cells. Hence this study presents an example of a pathological change of cytokeratin filaments and illustrates the value of monoclonal antibodies in the detection of such changes.