Li Qian, Huo Yongwei, Guo Yonghong, Zheng Xiaoyan, Sun Wengang, Hao Zhiming
Department of Rheumatology, the First Affiliated Hospital of Xi'an Jiaotong University, Xi'an 710061, China.
Research Center of Reproductive Medicine, Medical School of Xi'an Jiaotong University, Xi'an 710061, China.
Molecules. 2017 Apr 28;22(5):706. doi: 10.3390/molecules22050706.
Gremlin-1, a highly conserved glycosylated and phosphorylated secretory protein, plays important roles in diverse biological processes including early embryonic development, fibrosis, tumorigenesis, and renal pathophysiology. Aptamers, which are RNA or DNA single-stranded oligonucleotides capable of binding specifically to different targets ranging from small organics to whole cells, have potential applications in targeted imaging, diagnosis and therapy. In this study, we obtained a DNA aptamer against Gremlin-1 (G-ap49) using in vitro Systematic Evolution of Ligands by Exponential Enrichment (SELEX). Binding assay and dot-blot showed that G-ap49 had high affinity for Gremlin-1. Further experiments indicated that G-ap49 was quite stable in a cell culture system and could be used in South-Western blot analysis, enzyme-linked aptamer sorbent assay (ELASA), and aptamer-based cytochemistry and histochemistry staining to detect Gremlin-1. Moreover, our study demonstrated that G-ap49 is capable of revealing the subcellular localization of Gremlin-1. These data indicate that G-ap49 can be used as an alternative to antibodies in detecting Gremlin-1.
Gremlin-1是一种高度保守的糖基化和磷酸化分泌蛋白，在多种生物学过程中发挥重要作用，包括早期胚胎发育、纤维化、肿瘤发生和肾脏病理生理学。适体是能够特异性结合从小分子有机物到全细胞等不同靶标的RNA或DNA单链寡核苷酸，在靶向成像、诊断和治疗方面具有潜在应用。在本研究中，我们使用指数富集的配体系统进化技术（SELEX）体外筛选获得了一种针对Gremlin-1的DNA适体（G-ap49）。结合实验和斑点印迹显示G-ap49对Gremlin-1具有高亲和力。进一步实验表明，G-ap49在细胞培养系统中相当稳定，可用于蛋白质印迹分析、酶联适体吸附测定（ELASA）以及基于适体的细胞化学和组织化学染色以检测Gremlin-1。此外，我们的研究表明G-ap49能够揭示Gremlin-1的亚细胞定位。这些数据表明，G-ap49可作为检测Gremlin-1时替代抗体的一种选择。
