Ligand-guided selection of aptamers against T-cell Receptor-cluster of differentiation 3 (TCR-CD3) expressed on Jurkat.E6 cells.
Ligand-guided selection of aptamers against T-cell Receptor-cluster of differentiation 3 (TCR-CD3) expressed on Jurkat.E6 cells.
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DOI：
10.1016/j.ab.2016.08.007
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发表时间：
2016-11-01
影响因子：
2.9
通讯作者：
Mallikaratchy PR
中科院分区：
文献类型：
--
作者：
Zumrut HE;Ara MN;Maio GE;Van NA;Batool S;Mallikaratchy PR
我们最近引入了一种筛选技术，称为配体引导选择，（LIGS），选择性地确定目标特异性适配子从进化的细胞-SELEX库。Cell-SELEX利用大型组合单链寡核苷酸文库，并通过重复的分区和扩增循环，逐步选择具有可变DNA结合亲和力和特异性的针对全细胞的DNA配体。LIGS利用分配步骤并引入第二个预先存在的高亲和力单克隆抗体（mAb）配体，以击败并洗脱特定的适体，使其与抗体而不是细胞的结合靶点结合。在这里，使用针对分化簇3（CD 3 ε）的抗CD 3 ε mAb作为针对Jurkat.E6细胞上表达的T细胞受体（TCR）复合物的一个结构域的指导配体，我们发现了针对人T淋巴细胞永生化系上表达的TCR复合物的三种特异性适体。总之，我们证明了可以利用针对多结构域蛋白复合物的单个结构域的抗体在其内源状态下鉴定特异性适体，而不需要后或前SELEX蛋白质操作。
We recently introduced a screening technology termed ligand-guided selection, (LIGS), to selectively identify target-specific aptamers from an evolved cell-SELEX library. Cell-SELEX utilizes a large combinatorial single-stranded oligonucleotide library and progressively selects DNA ligands against whole cells with variable DNA-binding affinities and specificities by repeated rounds of partition and amplification. LIGS exploits the partition step and introduces a secondary, pre-existing high-affinity monoclonal antibody (mAb) ligand to outcompete and elute specific aptamers towards the binding target of the antibody, not the cell. Here, using anti-CD3ε mAb against the cluster of differentiation 3 (CD3ε), as the guiding ligand against one of the domains of the T-Cell Receptor (TCR) complex expressed on Jurkat.E6 cells, we discovered three specific aptamers against TCR complex expressed on an immortalized line of human T lymphocyte cells. In sum, we demonstrate that specific aptamers can be identified utilizing an antibody against a single domain of a multidomain protein complex in their endogenous state with neither post- nor pre-SELEX protein manipulation.
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