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利用核糖体展示技术筛选针对ESAT-6抗原的单链可变片段

Selection of single-chain variable fragments specific for ESAT-6 antigen using ribosome display.

作者信息

Ahangarzadeh Shahrzad, Bandehpour Mojgan, Kazemi Bahram

机构信息

Department of Biotechnology, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran.

Cellular and Molecular Biology Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran; Department of Biotechnology, School of Advanced Technologies in Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran.

出版信息

Iran J Basic Med Sci. 2017 Mar;20(3):327-333. doi: 10.22038/ijbms.2017.8363.

DOI:10.22038/ijbms.2017.8363
PMID:28392906
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC5378971/
Abstract

OBJECTIVES

Tuberculosis (TB) is still one of the problematic infectious diseases in developing countries, especially in Iran. In the present study, we applied ribosome display technique to select single chain variable fragments (scFvs) specific for the 6-kDa early secretory antigenic target (ESAT-6) antigen of from a mouse scFv library.

MATERIALS AND METHODS

The gene encoding ESAT-6 was cloned into pET22b(+) plasmid and expressed in BL21 (DE3). The purified recombinant ESAT-6 protein was injected into female BALB/c mice for immunization, and then m-RNA was extracted from the spleen of immunized mice. The anti-ESAT-6 VH/k chain library was assembled by joining of VH and k into the VH/k chain with a 72-bp DNA linker by SOE (splicing by overlap extension) PCR. The scFv library was panned against ESAT-6 using a single round of ribosome display via a rabbit reticulocyte lysate system.

RESULTS

ELISA assay showed that one of the selected scFvs had higher affinity against the recombinant ESAT-6 protein. The affinity of the candidate scFv was ~ 3.74×10 M.

CONCLUSION

It could be proposed that the isolated scFv in this study may be useful for the diagnosis of TB.

摘要

目的

结核病（TB）仍是发展中国家尤其是伊朗存在问题的传染病之一。在本研究中，我们应用核糖体展示技术从鼠源单链抗体可变区（scFv）文库中筛选针对结核分枝杆菌6 kDa早期分泌性抗原靶标（ESAT-6）抗原的scFv。

材料与方法

将编码ESAT-6的基因克隆到pET22b(+)质粒中，并在大肠杆菌BL21(DE3)中表达。将纯化的重组ESAT-6蛋白注射到雌性BALB/c小鼠中进行免疫，然后从免疫小鼠的脾脏中提取mRNA。通过重叠延伸PCR（SOE）将重链可变区（VH）和轻链可变区（k）与72 bp的DNA接头连接，构建抗ESAT-6 VH/k链文库。通过兔网织红细胞裂解液系统，利用一轮核糖体展示技术对scFv文库进行针对ESAT-6 的淘选。

结果

ELISA分析表明，筛选出的一种scFv对重组ESAT-6蛋白具有更高的亲和力。候选scFv的亲和力约为3.74×10 M。

结论

可以认为本研究中分离出的scFv可能对结核病的诊断有用。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ffc/5378971/acbfa674c28a/IJBMS-20-327-g006.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ffc/5378971/c34ef823ebd9/IJBMS-20-327-g001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ffc/5378971/168eb34744c6/IJBMS-20-327-g002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ffc/5378971/a3cbfcc5d95b/IJBMS-20-327-g003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ffc/5378971/9f1139fbb56f/IJBMS-20-327-g004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ffc/5378971/367281a59c64/IJBMS-20-327-g005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ffc/5378971/acbfa674c28a/IJBMS-20-327-g006.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ffc/5378971/c34ef823ebd9/IJBMS-20-327-g001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ffc/5378971/168eb34744c6/IJBMS-20-327-g002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ffc/5378971/a3cbfcc5d95b/IJBMS-20-327-g003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ffc/5378971/9f1139fbb56f/IJBMS-20-327-g004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ffc/5378971/367281a59c64/IJBMS-20-327-g005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ffc/5378971/acbfa674c28a/IJBMS-20-327-g006.jpg

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2
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Iran J Microbiol. 2016 Feb;8(1):29-35.
3
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4
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Front Microbiol. 2022 Nov 29;13:1043214. doi: 10.3389/fmicb.2022.1043214. eCollection 2022.
5
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Iran J Biotechnol. 2021 Jan 1;19(1):e2617. doi: 10.30498/IJB.2021.2617. eCollection 2021 Jan.
6
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7
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