A Highly Sensitive Detection System based on Proximity-dependent Hybridization with Computer-aided Affinity Maturation of a scFv Antibody.
A Highly Sensitive Detection System based on Proximity-dependent Hybridization with Computer-aided Affinity Maturation of a scFv Antibody.
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基于邻近依赖性杂交和计算机辅助 scFv 抗体亲和力成熟的高灵敏检测系统。
DOI：
10.1038/s41598-018-22111-4
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发表时间：
2018-03-01
影响因子：
4.6
通讯作者：
Chen A
中科院分区：
文献类型：
--
作者：
Wang Z;Li Y;Liang W;Zheng J;Li S;Hu C;Chen A
乙型肝炎病毒（HBV）感染是世界范围内的一个严重健康问题，而HBV preS1是监测HBV感染的重要生物标志物。此前，我们发现鼠单克隆抗体 mAb-D8 靶向 HBV 的 preS1 (aa91-107) 片段。为了提高其性能，我们制备了mAb-D8（scFvD8）的单链可变区，并通过计算机建模构建了scFvD8-preS1（aa91-107）复合物的三维结构。通过引入突变L96Tyr至Ser和H98Asp至Ser，scFvD8的亲和力显着增加。此外，基于邻近依赖性杂交策略设计了一种高灵敏度免疫传感器，其中 preS1 抗原与 DNA 标记的抗体竞争性反应，导致邻近依赖性杂交减少，Fc 片段的电化学信号增加，可用于 preS1 的定量。结果显示检测范围较宽，从 1 pM 到 50 pM，检测限为 0.1 pM。该免疫传感器在临床血清样本中的灵敏度和特异性分别为100%和96%。这项研究提供了一种基于邻近依赖性杂交和 scFv 抗体片段的新型系统，用于高灵敏度地快速定量感兴趣的抗原。
The hepatitis B virus (HBV) infection is a critical health problem worldwide, and HBV preS1 is an important biomarker for monitoring HBV infection. Previously, we found that a murine monoclonal antibody, mAb-D8, targets the preS1 (aa91-107) fragment of HBV. To improve its performance, we prepared the single-chain variable region of mAb-D8 (scFvD8) and constructed the three-dimensional structure of the scFvD8-preS1 (aa91-107) complex by computer modelling. The affinity of scFvD8 was markedly increased by the introduction of mutations L96Tyr to Ser and H98Asp to Ser. Furthermore, a highly sensitive immunosensor was designed based on a proximity-dependent hybridization strategy in which the preS1 antigen competitively reacts with an antibody labelled with DNA, resulting in decreased proximity-dependent hybridization and increased electrochemical signal from the Fc fragment, which can be used for the quantisation of preS1. The results showed a wide detection range from 1 pM to 50 pM with a detection limit of 0.1 pM. The sensitivity and specificity of this immunosensor in clinical serum samples were 100% and 96%, respectively. This study provides a novel system based on proximity-dependent hybridization and the scFv antibody fragment for the rapid quantisation of antigens of interest with a high sensitivity.
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影响因子：
5
作者：
Ahangar, Laleh Enayati;Mehrgardi, Masoud A.
通讯作者：
Mehrgardi, Masoud A.
影响因子：
4.4
作者：
Hur, Byung-ung;Choi, Hyo-jung;Cha, Sang-hoon
通讯作者：
Cha, Sang-hoon
基于多色量子点和金纳米粒子的多重电化学发光DNA传感器测定乙型肝炎病毒和丙型肝炎病毒
影响因子：
6.2
作者：
Liu, Linlin;Wang, Xinyan;Su, Xingguang
通讯作者：
Su, Xingguang
影响因子：
5.8
作者：
Nguyen, Minh N.;Pradhan, Mohan R.;Zhong, Pingyu
通讯作者：
Zhong, Pingyu
影响因子：
2.4
作者：
Heselpoth, Ryan D.;Yin, Yizhou;Nelson, Daniel C.
通讯作者：
Nelson, Daniel C.