Development and Preliminary Evaluation of a Serum Blocking ELISA for the Detection of PEDV Antibodies and Assessment of Its Association with Neutralizing Antibody Titers

Porcine epidemic diarrhea virus (PEDV), the etiological agent of porcine epidemic diarrhea (PED), remains a major threat to swine production worldwide. The genetic variation and the co-circulation of multiple genotypes of this virus may compromise vaccine-induced protection, creating an urgent need for practical serological tools for herd-level monitoring. A blocking enzyme-linked immunosorbent assay (blocking ELISA) was developed using a recombinant truncated PEDV S1 protein as the coating antigen and a horseradish peroxidase (HRP)-labeled anti-PEDV S1 monoclonal antibody as the competing detector. No cross-reactivity was detected with the tested antisera against porcine reproductive and respiratory syndrome virus (PRRSV), pseudorabies virus (PRV), classical swine fever virus (CSFV), or porcine circovirus type 2 (PCV2). The intra-assay and inter-assay coefficients of variation were 3.02% and 3.76%, respectively. Receiver operating characteristic analysis yielded an optimal cutoff of 31.76%, with a sensitivity of 98% and a specificity of 99%. Among 33 clinical serum samples, blocking-ELISA classifications were fully concordant with infected-cell indirect immunofluorescence assay (IFA) results (33/33, 100%), which were interpreted as a measure of general PEDV seroreactivity. Comparison with virus neutralization testing of 98 sera showed category-specific concordance rates of 81.25%, 90.91%, and 100% for neutralizing antibody titers of 1:64, 1:128, and ≥1:256, respectively. All sera in the ≤1:16 category were negative by blocking ELISA. A continuous correlation analysis between the blocking percentage and neutralizing antibody titer was performed (ρ = 0.484, 95% CI 0.315–0.623, p = 5.1 × 10−7; n = 97). Compared to traditional neutralization tests, this method features simpler operation, faster detection, and higher efficiency, rendering it suitable for large-scale monitoring of vaccine efficacy in swine herds and offering robust technical support for precise evaluation of PEDV immunity.