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The Thai Journal of Veterinary Medicine

Abstract

Newcastle disease (ND) is an avian respiratory disease caused by the Newcastle disease virus (NDV) and is prevalent worldwide but lacks a reliable treatment to surmount it. The single-chain fragment variable (scFv) antibody is the smallest functional unit in an immunoglobulin retaining antigen-binding activities. Previous studies have demonstrated that by acting against the NDV phosphoprotein, scFv is capable of neutralizing the virus in infected cells. In this study, two anti-NDV scFv expressing cassettes driven by CAG promoter and EF1α promoter, respectively, were constructed using the piggyBac transposon plasmid backbone. The recombinant plasmids were transfected into DF-1 and 293T cells and four stable cell lines CAG-scFv-DF-1, EF1α-scFv-DF-1, CAG-scFv-293T and EF1α-scFv-293T were established after puromycin screening. RT-PCR results confirmed the presence of anti-NDV scFv mRNA in all four cell lines. Western blot results showed that scFv was detected in the culture supernatant of the CAG-scFv-DF-1 and EF1α-scFv-DF-1 lines, indicating that the scFv antibody was secretory after expression in the cells. Cytopathic effect assay showed that cells expressing scFv were more resistant to NDV F48E9 than those of control when the virus titer was not higher than 60×TCID50. Results in this study offer information to the generation of transgenic chickens resistant to NDV infection and for production of neutralizing antibodies against NDV.

Publisher

Faculty of Veterinary Science, Chulalongkorn University

First Page

293

Last Page

301

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