Development of ELISA Using Phage-Displayed Stx2 Mini-Body for Detection of STEC Antigen in Field Farming Pig Samples

Porcine edema disease (ED), which causes enormous economic losses in pig farms, is caused by Shiga toxin type 2e (Stx2e) Escherichia coli (STEC), which frequently occurs in young piglets. In this study, we aimed to express a fused Stx2e peptide on a phage surface to generate an innovative sandwich ELISA for the detection of STEC antigen in field pig farming samples. The amino acid sequences at positions 241-319 were selected for capture antibody (T1D2) production. T1D2 was selected after the third round of biopanning, and it showed a high yield with no major impurities. T1D2-ELISA can detect recombinant modified Stx2e antigen, and the detection limit of the antigen was approximately below 20 pg/mL. The sensitivity of T1D2-ELISA was determined using five different stool samples, with a total of 25 stool samples. Positive Stx2e antigen samples were detected only in one of the 25 samples using T1D2-ELISA. The ELISA values of positive stool samples were >300 pg and <600 pg. In conclusion, we developed an innovative ELISA for the detection of STEC antigens in field pig farming samples. It can also be used to easily detect STEC antigens in porcine stool samples. We anticipate that our novel T1D2-ELISA method will enable the effective monitoring of STEC antigen content during industrial vaccine production. By leveraging this approach, we aimed to enhance production efficiency and ensure high-quality vaccines.