Cloning, Expression and Functionality Evaluation of Recombinant Monoclonal Antibody Against VP1 Capsid Protein of FMD Virus

Document Type : Original Articles

Authors

1 Department of Pathobiology, SR.C., Islamic Azad University, Tehran, Iran.

2 Department of Human Viral Vaccines, Razi Vaccine and Serum Research Institute (RVSRI), Agricultural Research, Education and Extension Organization (AREEO), Karaj, Iran.

3 Department of Microbiology, Ka.C., Islamic Azad University, Karaj, Iran.

10.32598/ARI.81.1.3491

Abstract

Introduction: Foot-and-mouth disease (FMD) is a highly contagious viral disease caused by the FMD virus (FMDV), which belongs to the Aphthovirus genus in the Picornaviridae family. FMDV is a highly variable RNA virus, and there is limited cross-protection between serotypes and strains. The disease can have devastating economic, social, and environmental impacts. The FMDV capsid is composed of VP1-VP4 structural proteins, with VP1 being the most abundant protein consisting of 213 amino acids. The G-H loop (residues 141-160) of the VP1 capsid protein is highly variable and serves as the main antigenic region. It contains a conserved triplet of amino acids, Arg-Gly-Asp (RGD), which induces the production of protective antibodies against various FMDV types. Monoclonal antibodies (mAbs) play a crucial role in detecting and serotyping FMDV in pathological samples, as well as evaluating protection post-vaccination against FMD.
Materials & Methods: This study explores the expression and function of an engineered recombinant single-chain variable fragment (scFv-mAb) in Escherichia coli BL21 (DE3) Rosetta strain as a cost-effective prokaryotic system with high yield. The scFv-mAb gene was inserted into the pET28a (+) expression plasmid. E. coli cells were transformed with the plasmid, induced with 0.5 mM IPTG, and incubated at 37 ˚C for 12 hours. The protein was purified using a Ni2+-NTA resin column and analyzed by 12% SDS-PAGE for quality assessment. The efficiency and functionality of the scFv-mAb were confirmed using an indirect sandwich (capture) enzyme-linked immunosorbent assay (ELISA).
Results: The purified scFv-mAb concentration was determined to be approximately 2.00 mg/mL using the Bradford assay under optimal conditions. Each well was coated with 400 ng of scFv-mAb based on checkerboard results and the mean of negative serum at a 1:10 dilution. The indirect sandwich ELISA assay yielded an optical density (OD) signal range of 0.3 to 1.5 at a 450 nm wavelength in different positive control treatments.
Conclusion: The ELISA results showed that the scFv-mAb fragment successfully detected serotype O of FMDV. Further research could confirm the potential of this recombinant antibody for broader commercial applications in the future.

Keywords

Archives of Razi Institute
Volume 81, Issue 1
January and February 2026
Pages 147-156

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