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Expression and purification of an anti-clenbuterol single chain Fv antibody in Escherichia coli.
Protein Expr Purif. 2010 Jul; 72(1):26-31.PE

Abstract

Recombinant antibodies with desirable characteristics that can replace polyclonal or monoclonal antibodies are important for enzyme-linked immunosorbent assay (ELISA) of residues of clenbuterol (CBL), an illicit veterinary drug. Here, we report our work on expression and purification of a mouse-derived anti-CBL single chain Fv (scFv) antibody in Escherichia coli (E. coli). An expression plasmid pBV220-CBL was constructed and transformed into E. coli BL21 (DH3) strain cells. After induction by temperature, the 6x His-tagged anti-CBL scFv antibodies were expressed with the yield of 31%. The solubilized inclusion bodies were extracted, denatured and then purified by Ni-NTA column chromatography. The purified recombinant target protein was analyzed by high performance liquid chromatography, SDS-PAGE and Western blotting, respectively. The results showed the prepared anti-CBL scFv antibodies posed HRP-anti-His-tag antibody-recognized activity and their purity was up to 96%. Moreover, an indirect competitive ELISA based on the anti-CBL scFv antibodies revealed that the limit of detection for CBL was 0.5 ng/ml and the linear range was 1.5-10.6 ng/ml. Taken together, these findings suggest that the prepared recombinant antibody can be used for future immunoassay detection for CBL.

Authors+Show Affiliations

Key Laboratory of Food Quality and Safety of Guangdong Province/College of Food Science, South China Agricultural University, Guangzhou 510642, PR China.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

20206697

Citation

Wang, Hong, et al. "Expression and Purification of an Anti-clenbuterol Single Chain Fv Antibody in Escherichia Coli." Protein Expression and Purification, vol. 72, no. 1, 2010, pp. 26-31.
Wang H, Liu X, He Y, et al. Expression and purification of an anti-clenbuterol single chain Fv antibody in Escherichia coli. Protein Expr Purif. 2010;72(1):26-31.
Wang, H., Liu, X., He, Y., Dong, J., Sun, Y., Liang, Y., Yang, J., Lei, H., Shen, Y., & Xu, X. (2010). Expression and purification of an anti-clenbuterol single chain Fv antibody in Escherichia coli. Protein Expression and Purification, 72(1), 26-31. https://doi.org/10.1016/j.pep.2010.03.001
Wang H, et al. Expression and Purification of an Anti-clenbuterol Single Chain Fv Antibody in Escherichia Coli. Protein Expr Purif. 2010;72(1):26-31. PubMed PMID: 20206697.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Expression and purification of an anti-clenbuterol single chain Fv antibody in Escherichia coli. AU - Wang,Hong, AU - Liu,Xixia, AU - He,Yongsheng, AU - Dong,Jiexian, AU - Sun,Yuanming, AU - Liang,Yan, AU - Yang,Jinyi, AU - Lei,Hongtao, AU - Shen,Yudong, AU - Xu,Xiaoyan, Y1 - 2010/03/04/ PY - 2009/12/03/received PY - 2010/02/23/revised PY - 2010/03/01/accepted PY - 2010/3/9/entrez PY - 2010/3/9/pubmed PY - 2010/7/16/medline SP - 26 EP - 31 JF - Protein expression and purification JO - Protein Expr Purif VL - 72 IS - 1 N2 - Recombinant antibodies with desirable characteristics that can replace polyclonal or monoclonal antibodies are important for enzyme-linked immunosorbent assay (ELISA) of residues of clenbuterol (CBL), an illicit veterinary drug. Here, we report our work on expression and purification of a mouse-derived anti-CBL single chain Fv (scFv) antibody in Escherichia coli (E. coli). An expression plasmid pBV220-CBL was constructed and transformed into E. coli BL21 (DH3) strain cells. After induction by temperature, the 6x His-tagged anti-CBL scFv antibodies were expressed with the yield of 31%. The solubilized inclusion bodies were extracted, denatured and then purified by Ni-NTA column chromatography. The purified recombinant target protein was analyzed by high performance liquid chromatography, SDS-PAGE and Western blotting, respectively. The results showed the prepared anti-CBL scFv antibodies posed HRP-anti-His-tag antibody-recognized activity and their purity was up to 96%. Moreover, an indirect competitive ELISA based on the anti-CBL scFv antibodies revealed that the limit of detection for CBL was 0.5 ng/ml and the linear range was 1.5-10.6 ng/ml. Taken together, these findings suggest that the prepared recombinant antibody can be used for future immunoassay detection for CBL. SN - 1096-0279 UR - https://www.unboundmedicine.com/medline/citation/20206697/Expression_and_purification_of_an_anti_clenbuterol_single_chain_Fv_antibody_in_Escherichia_coli_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S1046-5928(10)00064-1 DB - PRIME DP - Unbound Medicine ER -