Production and application of recombinant antibodies to foot-and-mouth disease virus non-structural protein 3ABC.

摘要

The stamping out of animals to control a foot-and-mouth disease (FMD) outbreak results in enormous livestock losses. The implementation of vaccination strategies can reduce these losses; however it complicates the process of establishing freedom from disease following an outbreak. The availability of quality diagnostic tests to differentiate infected from vaccinated animals (DIVA) is crucial to prove freedom from disease and allow for the resumption of trade in livestock products. All current foot-and-mouth disease virus (FMDV) DIVA tests rely on polyclonal or monoclonal hybridoma derived antibody reagents, which can be difficult to prepare and maintain in a quality-assured manner and in the quantities required for post-outbreak surveillance. Recombinant antibodies can be produced in large quantities at low cost in bacteria to guarantee the supply of a consistent and well-characterised reagent. The production of recombinant antibodies does not rely on animal immunisation and does not require the maintenance of viable hybridoma cell lines. In this study, phage display libraries of recombinant antibody single chain variable fragments (scFv) against FMDV were generated from chickens immunised with recombinant non-structural protein (NSP) 3ABC. A total of 32 positive clones were obtained that represented three distinctive genetic sequences, Chicken Recombinant Antibody-Foot-and-Mouth disease (CRAb-FM) 26, -FM27 and -FM29. Each was shown to bind the 3B region of the 3ABC protein. When evaluated in a C-ELISA format using sera derived from cattle, sheep and pigs representing naive, FMDV-vaccinated or FMDV-infected animals, CRAb-FM27 gave the best performance when paired with an E. coli-derived recombinant 3ABC, demonstrating the potential to be used as a species- and serotype-independent FMDV DIVA test. To our knowledge, this is the first FMDV DIVA test that uses both recombinant antibody and antigen derived from bacterial expression systems.