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Issue 30, 2017
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Production and evolution of a ScFv antibody for immunoassay of residual phenothiazine drugs in meat based on computational simulation

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Abstract

In this study, the gene of a single chain variable fragment (ScFv) from a hybridoma cell strain excreting the monoclonal antibody for 2-chlorophenothiazine was directly transformed into E. coli to express the ScFv antibody. The obtained ScFv antibody showed similar recognition performances for 5 phenothiazine drugs to its parental monoclonal antibody. Its molecular recognition mechanisms for the 5 drugs were studied by using molecular docking, and then the ScFv antibody was evolved directionally to generate a ScFv mutant by mutagenesis of a contact amino acid Phe164 to Pro based on the analysis of virtual mutation. The molecular docking showed that both the mutant-analyte intermolecular forces and the total binding energies increased, so the mutant showed highly improved sensitivity to the 5 drugs with up to 13 fold decreased IC50 values. Then an indirect competitive immunoassay was developed to determine the residues of the 5 drugs in meat. The limits of detection were in the range of 0.1–1.8 ng g−1, and the recoveries from the fortified blank meat sample were in the range of 66.4–97.2%. Furthermore, the results of the immunoassay of the real samples were consistent with those of a high performance liquid chromatography method.

Graphical abstract: Production and evolution of a ScFv antibody for immunoassay of residual phenothiazine drugs in meat based on computational simulation

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Publication details

The article was received on 28 Apr 2017, accepted on 04 Jul 2017 and first published on 04 Jul 2017


Article type: Paper
DOI: 10.1039/C7AY01103B
Citation: Anal. Methods, 2017,9, 4455-4463
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    Production and evolution of a ScFv antibody for immunoassay of residual phenothiazine drugs in meat based on computational simulation

    F. S. Shi, L. Zhang, W. Q. Xia, J. Liu, H. C. Zhang and J. P. Wang, Anal. Methods, 2017, 9, 4455
    DOI: 10.1039/C7AY01103B

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