PDF(Pubmed)
Abstract：
Nanobodies (Nbs) are recombinant single-domain fragments derived from camelids\' heavy-chain antibodies (HCAbs). Nanobodies are increasingly used in numerous biotechnological and medical applications because of their high stability, solubility, and yield. However, one major obstacle prohibiting Nb expansion is the affordability of specific detector antibodies for their final revelation. In this work, the production of a specific anti-Nb antibody as a general detector for camel antibodies, conventional cIgG, and HCAb, and their derived Nbs was sought. Thus, a T7 promoter plasmid was constructed and used to highly express six different Nbs that were used in a successful rabbit immunization. Affinity-purified rabbit anti-Nb rIgG was able to detect immobilized or antigen-bound Nbs via enzyme-linked immunosorbent assay, and its performance was comparable to that of a commercial anti-6× His antibody. Its capacities in dosing impure Nbs, detecting Nbs displayed on M13 phages, and revealing denatured Nbs in immune blotting were all proven. As expected, and because of shared epitopes, rabbit anti-Nb cross-reacted with cIgG, HCAbs, and 6× His-tagged proteins, and the percentage of each fraction within anti-Nb rIgG was determined. Anti-Nb is a promising tool for the checkpoints throughout the recombinant Nb technology.
摘要：
纳米抗体(Nbs)是源自骆驼科动物重链抗体(HCAb)的重组单结构域片段。纳米抗体由于其高稳定性而越来越多地用于许多生物技术和医学应用。溶解度，和产量。然而,阻止Nb膨胀的一个主要障碍是特异性检测抗体对其最终启示的可负担性。在这项工作中,产生特异性抗Nb抗体作为骆驼抗体的通用检测器，常规cIgG，和HCAb,并寻求他们派生的NBS。因此，构建T7启动子质粒并用于高度表达在成功的兔免疫中使用的六种不同的Nbs。亲和纯化的兔抗NbrIgG能够通过酶联免疫吸附试验检测固定或抗原结合的Nbs，其性能与商业抗6×His抗体相当。它的能力在剂量不纯的NBS，检测M13噬菌体上显示的NBS，并在免疫印迹中揭示变性的Nbs都得到了证实。不出所料,因为共有的表位，兔抗Nb与cIgG交叉反应，HCABs,和6×His标记的蛋白质，并确定抗NbrIgG中每个部分的百分比。反Nb是整个重组Nb技术中用于检查点的有前途的工具。
