Cui He, Lan Xi, Lu Shemin, Zhang Fujun, Zhang Wanggang
Department of Clinical Hematology, The Affiliated Hospital, Inner Mongolia Medical University, No. 1 Tongdao North Street, Hohhot 010059, China.
Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Xi'an Jiaotong University Health Science Center, No. 76 The Yanta West Road, Xi'an, Shaanxi 710061, China.
J Pharm Anal. 2017 Jun;7(3):190-195. doi: 10.1016/j.jpha.2017.02.001. Epub 2017 Feb 16.
Monoclonal antibodies (MAbs) are important tools for the study of proteins' function and structure. But there has been no report on the preparation of MAbs against human KIAA0100 protein up to date. Here, first, we generated the mouse MAb against human KIAA0100 protein using purified recombinant 6×Histidinc (6×His)-tagged human KIAA0100 protein segment (1557-2234) as an antigen; then, the mRNA expression of human KIAA0100 gene was detected in U937 cells using Northern blot analysis. The results showed that the mouse MAb against human KIAA0100 protein could sensitively recognize the human KIAA0100 protein using Western blot analysis and immunocytochemistry analysis. Besides, Western blot analysis revealed that human KIAA0100 gene possibly encoded two different protein products (254 kDa and <250 kDa) in U937 cells. Moreover, Northern blot analysis confirmed that human KIAA0100 gene might produced two different mRNA products (6000-10000 bp and 5000-6000 bp) in U937 cells. The results provide a basis for large-scale production of the MAb against human KIAA0100 protein, which will be useful for the study of human KIAA0100 protein's function/structure and MAb-targeted drugs in the future.
单克隆抗体（MAb）是研究蛋白质功能和结构的重要工具。但迄今为止，尚无关于制备抗人KIAA0100蛋白单克隆抗体的报道。在此，首先，我们以纯化的重组6×组氨酸（6×His）标签的人KIAA0100蛋白片段（1557 - 2234）作为抗原，制备了抗人KIAA0100蛋白的小鼠单克隆抗体；然后，使用Northern印迹分析检测了U937细胞中人KIAA0100基因的mRNA表达。结果表明，通过蛋白质印迹分析和免疫细胞化学分析，抗人KIAA0100蛋白的小鼠单克隆抗体能够灵敏地识别该蛋白。此外，蛋白质印迹分析显示，人KIAA0100基因在U937细胞中可能编码两种不同的蛋白质产物（254 kDa和<250 kDa）。而且，Northern印迹分析证实，人KIAA0100基因在U937细胞中可能产生两种不同的mRNA产物（6000 - 10000 bp和5000 - 6000 bp）。这些结果为大规模生产抗人KIAA0100蛋白的单克隆抗体提供了依据，这将有助于未来对人KIAA0100蛋白的功能/结构以及单克隆抗体靶向药物的研究。
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