Application of Monoclonal Antibodies Against Mouse Dermokine.
Application of Monoclonal Antibodies Against Mouse Dermokine.
复制标题
抗小鼠皮肤因子单克隆抗体的应用。
DOI：
10.1089/mab.2016.0051
复制
发表时间：
2017
影响因子：
--
通讯作者：
Tachibana T.
中科院分区：
文献类型：
--
作者：
Higashi K;Wang CX;Yokoyama C;Yamada K;Saito K;Tachibana T.
Dermokine是角质形成细胞分化过程中最高表达的蛋白质之一。据报道，小鼠皮肤因子由22个外显子编码，其表达导致三种转录物β、γ和α，它们从两个不同的转录起始位点转录。α亚型代表β亚型的羧基末端结构域，而γ亚型缺乏该结构域。为了揭示每种亚型在小鼠中的分布和表达水平，我们制备了针对皮肤因子-β/γ和皮肤因子-β/α的大鼠单克隆抗体。在免疫荧光研究中，高水平的细胞外钙显著升高培养的小鼠角质形成细胞的胞质溶胶中的dermokine的表达水平。Western blot分析显示，高钙条件下，Dermokine-β和Dermokine-α的表达水平升高。最后，我们开发了一种基于单克隆抗体的敏感的夹心酶联免疫吸附试验（ELISA），并表明分泌到培养基中的皮肤因子-β从小鼠角质形成细胞显着增加的方式依赖于细胞外钙浓度。这些dermokine特异性抗体使我们能够获得新的见解的作用，每个dermokine亚型在皮肤的稳态。
Dermokine is one of the most highly expressed proteins in differentiating keratinocytes. Mouse dermokine has been reported to be encoded by 22 exons, and its expression leads to three transcripts, β, γ, and α, which are transcribed from two different transcriptional start sites. The α isoform represents the carboxyl-terminal domain of the β isoform, whereas the γ isoform lacks this domain. To reveal the distributions and expression levels of each isoform in mice, we generated rat monoclonal antibodies against dermokine-β/γ and dermokine-β/α. In immunofluorescence studies, the expression levels of dermokine in the cytosol of the cultured mouse keratinocytes were significantly elevated by high levels of extracellular calcium. In Western blot analyses, the expression levels of dermokine-β and dermokine-α were increased in the presence of high calcium. Finally, we developed a monoclonal antibody-based sensitive sandwich enzyme-linked immunosorbent assay (ELISA) and showed that the secreted dermokine-β into the culture medium from mouse keratinocytes was significantly increased in a manner dependent on the extracellular calcium concentration. These dermokine-specific antibodies have allowed us to gain new insights into the role of each dermokine isoform in cutaneous homeostasis.