A neutralizing nanobody-based liquid-phase blocking ELISA to assess the protective potency of Senecavirus A vaccine.

Senecavirus A (SVA) causes swine blister diseases in the crown of hooves, abortion syndrome, and increased mortality rates in piglets. Vaccination is the best defense against SVA infection. However, evaluations of the efficacy of SVA vaccines currently rely on challenging pigs with SVA, which is time-consuming, labor-intensive, and inconsistent with animal welfare mandates. To establish a replacement for the virus neutralization test (VNT) and SVA challenge tests, we developed a liquid-phase blocking enzyme-linked immunosorbent assay (nbLPB-ELISA) based on V1-VHH as the coating antibody (Ab) and biotinylated V1-VHH as the detection Ab. Under optimized conditions, the VNT and nbLPB-ELISA results were strongly correlated (Pearson R(2) = 0.84; p < 0.00001). Analysis of the LBP-ELISA and vaccine protection rate revealed that neutralizing Ab titers greater than 256 provided 100% protection, while titers of 64 and 128 offered 57.1% and 70% protection, respectively. The nbLPB-ELISA is a rapid, simple, safe, and cost-effective method of detecting SVA as a replacement for the SVA VNT and vaccine-challenge assays. KEY POINTS: • We evaluated a specific, high-affinity, and neutralizing nanobody-targeting SVA. • The developed nbLPB-ELISA can replace viral neutralization tests. • The nbLPB-ELISA is suitable to evaluate protective immunity.