Ph.D. Program in Molecular, Cellular and Developmental Biology, The Graduate Center of the City University of New York, New York, NY, USA.
Ph.D. Programs in Chemistry and Biochemistry, The Graduate Center of the City University of New York, New York, NY, USA.
Methods Mol Biol. 2023;2570:13-38. doi: 10.1007/978-1-0716-2695-5_2.
Oligonucleotide ligands (DNA, RNA, or XNA), also known as aptamers, are selected against various target molecules using an iterative, evolutionary process called systematic evolution of ligands by exponential enrichment (SELEX). To select aptamers against complex cell surface proteins in their native state, a variant of SELEX termed ligand-guided selection (LIGS) was recently introduced. The significance of LIGS is rooted in its strategy of exploiting the selection step in SELEX to identify highly specific aptamers against known cell surface markers. Thus, in LIGS, a higher-affinity secondary ligand, such as a monoclonal antibody (mAb) to a whole-cell bound to an evolved SELEX library, is introduced to outcompete sequences against the mAb targeting cell surface protein or induce a conformational switch to destabilize the aptamer-surface cell surface protein resulting in elution of the sequences. Here, we describe the detailed method of LIGS utilized in identifying aptamers against T-cell receptor cluster of differentiation three complex (TCR-CD3) expressed in human T-cells and T-cell leukemia.
寡核苷酸配体（DNA、RNA 或 XNA），也称为适体，是通过一种称为指数富集的配体系统进化（SELEX）的迭代、进化过程来针对各种靶分子进行选择的。为了在天然状态下选择针对复杂细胞表面蛋白的适体，最近引入了一种称为配体引导选择（LIGS）的 SELEX 变体。LIGS 的重要性源于其利用 SELEX 中的选择步骤来鉴定针对已知细胞表面标志物的高度特异性适体的策略。因此，在 LIGS 中，引入了一种高亲和力的二级配体，例如针对整个细胞结合的进化 SELEX 文库的单克隆抗体（mAb），以与针对细胞表面蛋白的 mAb 竞争序列或诱导构象转变以破坏适体-表面细胞表面蛋白，从而导致洗脱序列。在这里，我们描述了用于鉴定人 T 细胞和 T 细胞白血病中表达的 T 细胞受体簇分化三复合物（TCR-CD3）的适体的 LIGS 的详细方法。
