Takei Junko, Itai Shunsuke, Furusawa Yoshikazu, Yamada Shinji, Nakamura Takuro, Sano Masato, Harada Hiroyuki, Fukui Masato, Kaneko Mika K, Kato Yukinari
1 Department of Antibody Drug Development, Tohoku University Graduate School of Medicine, Sendai, Japan.
2 Department of Oral and Maxillofacial Surgery, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan.
Monoclon Antib Immunodiagn Immunother. 2019 Jun;38(3):129-132. doi: 10.1089/mab.2019.0012. Epub 2019 May 21.
Podoplanin (PDPN) is expressed on podocytes of the kidneys, type I alveolar cells of the lungs, and lymphatic endothelial cells. PDPN comprises three platelet aggregation-stimulating (PLAG) domains (PLAG1, PLAG2, and PLAG3) in the N-terminus and PLAG-like domains in the middle of the PDPN protein. We have previously reported on an anti-tiger PDPN (tigPDPN) monoclonal antibody (mAb), PMab-231, which was developed using the Cell-Based Immunization and Screening (CBIS) method. PMab-231 is very useful in flow cytometry, Western blotting, and immunohistochemical analyses; however, the binding epitope of PMab-231 remains to be elucidated. This study aimed to investigate the epitopes of PMab-231, which was developed by CBIS method, using enzyme-linked immunosorbent assay. The results revealed that the critical epitopes of PMab-231 are Glu29, Asp30, Asp31, Ile32, Met33, Thr34, Pro35, Gly36, and Glu38 of tigPDPN, which is corresponding to PLAG1/2. The findings of our study can be applied to the production of more functional anti-tigPDPN mAbs.
血小板结合蛋白（PDPN）在肾脏足细胞、肺的I型肺泡细胞和淋巴管内皮细胞中表达。PDPN在N端包含三个血小板聚集刺激（PLAG）结构域（PLAG1、PLAG2和PLAG3），在PDPN蛋白中部包含类PLAG结构域。我们之前报道过一种抗老虎血小板结合蛋白（tigPDPN）单克隆抗体（mAb）PMab - 231，它是使用基于细胞的免疫和筛选（CBIS）方法开发的。PMab - 231在流式细胞术、蛋白质印迹和免疫组织化学分析中非常有用；然而，PMab - 231的结合表位仍有待阐明。本研究旨在使用酶联免疫吸附测定法研究通过CBIS方法开发的PMab - 231的表位。结果显示，PMab - 231的关键表位是tigPDPN的Glu29、Asp30、Asp31、Ile32、Met33、Thr34、Pro35、Gly36和Glu38，它们对应于PLAG1/2。我们的研究结果可应用于生产更多功能性抗tigPDPN单克隆抗体。
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