The Use of Distinctive Monoclonal Antibodies in FMD VLP- and P1-Based Blocking ELISA for the Seromonitoring of Vaccinated Swine

Abstract

The serum neutralization (SN) test has been regarded as the “gold standard” for seroconversion following foot-and-mouth disease virus (FMDV) vaccination, although a high-level biosafety laboratory is necessary. ELISA is one alternative, and its format is constantly being improved. For instance, standard polyclonal antisera have been replaced by monoclonal antibodies (MAbs) for catching and detecting antibodies, and inactive viruses have been replaced by virus-like particles (VLPs). To the best of current knowledge, however, no researchers have evaluated the performances of different MAbs as tracers. In previous studies, we successfully identified site 1 and site 2 MAbs Q10E and P11A. In this study, following the established screening platform, the VLPs of putative escape mutants from sites 1 to 5 were expressed and used to demonstrate that S11B is a site 3 MAb. Additionally, the vulnerability of VLPs prompted us to assess another diagnostic antigen: unprocessed polyprotein P1. Therefore, we established and evaluated the performance of blocking ELISA (bELISA) systems based on VLPs and P1, pairing them with Q10E, P11A, S11B, and the non-neutralizing TSG MAb as tracers. The results indicated that the VLP paired with S11B demonstrated the highest correlation with the SN titers (R2 = 0.8071, n = 63). Excluding weakly positive serum samples (SN = 16–32, n = 14), the sensitivity and specificity were 95.65% and 96.15% (kappa = 0.92), respectively. Additionally, the P1 pairing with Q10E also demonstrated a high correlation (R2 = 0.768). We also discovered that these four antibodies had steric effects on one another to varying degrees, despite recognizing distinct antigenic sites. This finding indicated that MAbs as tracers could not accurately detect specific antibodies, possibly because MAbs are bulky compared to a protomeric unit. However, our results still provide convincing support for the application of two pairs of bELISA systems: VLP:S11B-HRP and P1:Q10E-HRP.

Mapping of foot-and-mouth disease virus antigenic sites recognized by single-domain antibodies reveals different 146S particle specific sites and particle flexibility

“…They were later ( 13 ) found to have escape mutations at VP2-72 (C9) and VP1-43 and 44 (C8), which corresponds to sites 2 and 3, respectively ( Figure 7 ). MAb S11B that binds type O site 3 also appears to be 146S specific ( 46 ). Thus, for type O, also two independent antigenic sites exist that are 146S specific, as we observed also for type A.…”

Serological Conversion through a Second Exposure to Inactivated Foot-and-Mouth Disease Virus Expressing the JC Epitope on the Viral Surface

“…We selected site 1 because its immunogenic epitope can be modified without disrupting the icosahedral shape of the FMDV; it is even possible to insert an amino acid sequence that protrudes from the surface of the viral capsid in a linear manner [ 14 ]. Functionally, the antigenic site 1 induces a protective immunity [ 15 , 16 , 17 ].…”