Construction of an immunized rabbit phage display antibody library for screening microcystin-LR high sensitive single-chain antibody
文献类型： 外文期刊
作者： Xu, Chongxin 1 ; Miao, Wenjie 1 ; He, Yan 3 ; Zu, Yao 1 ; Liu, Xiaoqin 4 ; Li, Jianhong 1 ; Liu, Xianjin 2 ;
作者机构： 1.Nanjing Normal Univ, Coll Life Sci, Nanjing 210023, Jiangsu, Peoples R China
2.Jiangsu Acad Agr Sci, Inst Food Safety & Nutr, Minist Sci & Technol, Lab Food Qual,Safety State Key Lab Cultivat Base, Nanjing 210014, Jiangsu, Peoples R China
3.Chinese Acad Agr Sci, Inst Food Sci & Technol, Beijing 100081, Peoples R China
4.Huaihua Vocat & Tech Coll, Huaihua 418007, Peoples R China
关键词： Microcystin; Polyclonal antibodies; Phage display antibody library; ScFv; Immunoassay
期刊名称：INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES （ 影响因子：6.953； 五年影响因子：6.737 ）
ISSN： 0141-8130
年卷期： 2019 年 123 卷
页码：
收录情况： SCI
摘要： Microcystin-LR (MC-LR) is one of the most common biotoxin that pollutes water and agricultural products. The study aims to obtain the high sensitive anti-MC-LR single-chain antibody (scFv) for detecting MC-LR. Here, a MC-LR-immunized rabbit phage display scFv library with its capacity of 3.26 x 10(9) CFU/mL was constructed and used for anti-MC-LR phage scFv particles screening. After four rounds of biopanning, 18 positives were isolated and identified successfully. The most positive scFv (RscFv3) was expressed in Escherichia coli HB2151 and purified with a concentration of 796.7 mu g/mL, and the purified anti-MC-LR polyclonal antibodies (PAbs) were 3.6 mg/mL. The PAbs and scFv based indirect competitive enzyme linked immunosorbent assay (IC-ELISAs) were established for MC-LR and its analogues, and the results showed they all had high sensitive for MC-LR with detection limits of 0.03 and 0.05 mu g/L, and had strong cross-reactivity for MC-RR, MC-WR and MC-YR, respectively. The average recovery rate was 91.9% with coefficient of variation <6.8% for scFv-based IC-ELISA to detect MC-LR spiked in water samples, which met the requirement of indoor testing. The present results indicate that we have obtained a high sensitive anti-MC-LR scFv by the MC-LR-immunized phage library construction and screening, and the scFv-based IC-ELISA can be used for monitoring MC-LR in water samples. (C) 2018 Published by Elsevier B.V.
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