Development of sandwich Enzyme-Linked Immunosorbent Assay for the detection of porcine epidemic diarrhea virus in fecal samples

Porcine epidemic diarrhea virus (PEDV) causes acute diarrhea and dehydration in new-born piglets with subsequent economic losses to swine industry. In the current study, gene encoding of 381aa-792aa spike protein (S1) with the main epitope relative to virus neutralization of PEDV was amplified by RT-PCR and inserted into vector pET-30A(+). The plasmid was transferred into Escherichia coli BL21 (DE3). Meanwhile, recombinant protein expression was induced by isopropyl-P-galactopyranoside (IPTG). After denaturation and renaturation of inclusion bodies, the Si protein was obtained by using purified recombinant S1 protein in immunized female BALB/c mice. Monoclonal antibodies (MAb) against Si protein, named 4C7 by hybridoma technique were gained successfully. The result showed that MAb can specifically respond to S1 protein and PEDV via ELISA, Western bolt and immunofluorescence assay methods. A sandwich ELISA (S-ELISA) was established by using the captured monoclonal antibodies 4C7. The sensitivity and specificity were compared between S-ELISA and RTPCR, which showed similar sensitivity and specificity. This work indicated that S-ELISA would be a significant tool alongside a specific diagnostic reagent for PEDV in future.