Development of an Indirect ELISA Based on the PEDV S Protein and Evaluation of Its Correlation With Neutralizing Antibody and Immune Protection.

Porcine epidemic diarrhea virus (PEDV) is an enteric coronavirus that infects pigs of all ages. Notably, neonatal piglets (≤7 days old) in naïve herds can experience case fatality rates approaching 100%, with hallmark clinical signs of vomiting, diarrhea, and dehydration, thereby representing a major threat to swine herd health. Serum PEDV-specific IgG serves as a key serological marker of vaccine-induced herd immunity. Accordingly, we developed an indirect ELISA (iELISA) using Chinese hamster ovary (CHO) cell-expressed recombinant PEDV spike (S) protein as the coating antigen. Checkerboard titration identified the optimal antigen-coating concentration and serum dilution as 1 μg/mL and 1:300, respectively. Based on sample-to-positive (S/P) ratios from 50 PEDV-negative sera, the diagnostic cutoff was set at 0.796, corresponding to the mean plus 3 standard deviation (SD) of the negative control group (n = 50). The intra- and interassay coefficients of variation (CVs) were both <6.30%, indicating good precision (repeatability) and reproducibility. Specificity testing showed no detectable cross-reactivity with sera positive for five common porcine viruses (porcine deltacoronavirus (PDCoV), transmissible gastroenteritis virus (TGEV), porcine group A rotavirus (PoRVA), porcine reproductive and respiratory syndrome virus (PRRSV), and African swine fever virus (ASFV)). Moreover, PEDV-specific IgG measured by iELISA correlated strongly with virus-neutralizing (VN) antibody titers (r = 0.95), and both metrics were associated with protective outcomes at the herd level. Collectively, this assay provides an effective tool for evaluating protective immunity following PEDV vaccination.