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Generation and Characterization of a scFv Antibody Against T3SS Needle of Vibrio parahaemolyticus

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Abstract

Vibrio parahaemolyticus, a halophilic gram-negative bacterium, is a food-borne pathogen that largely inhabits marine and estuarine environments, and poses a serious threat to human and animal health all over the world. The hollow “needle” channel, a specific assemble of T3SS which exists in most of gram-negative bacteria, plays a key role in the transition of virulence effectors to host cells. In this study, needle protein VP1694 was successfully expressed and purified, and the fusion protein Trx-VP1694 was used to immunize Balb/c mice. Subsequently, a phage single-chain fragment variable antibody (scFv) library was constructed, and a specific scFv against VP1694 named scFv-FA7 was screened by phage display panning. To further identify the characters of scFv, the soluble expression vector pACYC-scFv-skp was constructed and the soluble scFv was purified by Ni2+ affinity chromatography. ELISA analysis showed that the scFv-FA7 was specific to VP1694 antigen, and its affinity constant was 1.07 × 108 L/mol. These results offer a molecular basis to prevent and cure diseases by scFv, and also provide a new strategy for further research on virulence mechanism of T3SS in V. parahaemolyticus by scFv.

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Acknowledgments

This work was supported by the Program for New Century Excellent Talents in University (Grant NCET-10-0010), the Fujian Fund for Distinguished Young Scientists (Grant 2009J06008), the National Agricultural Achievements Transformation Fund (Grant 2011GB2C400012), and Agricultural Five-new Engineering Projects of Fujian Development and Reform Commission.

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Authors and Affiliations

  1. The Ministry of Education Key Laboratory of Biopesticide and Chemical Biology, and School of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou, 350002, China

    Rongzhi Wang, Sui Fang, Shuangshuang **ang, Sumei Ling, Jun Yuan & Shihua Wang

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  1. Rongzhi Wang
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  2. Sui Fang
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  3. Shuangshuang **ang
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  4. Sumei Ling
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Correspondence to Shihua Wang.

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Wang, R., Fang, S., **ang, S. et al. Generation and Characterization of a scFv Antibody Against T3SS Needle of Vibrio parahaemolyticus . Indian J Microbiol 54, 143–150 (2014). https://doi.org/10.1007/s12088-013-0428-6

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  • DOI: https://doi.org/10.1007/s12088-013-0428-6

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