Preparation of Ractopamine Single-Chain Variable Fragment and Development of icELISA Based on Immunomagnetic Beads

Ractopamine (RAC) is a feed additive that regulates protein synthesis. In this study, an indirect enzyme-linked immunosorbent assay (icELISA) for the detection of RAC residues in the porcine liver was developed based on our specific immunomagnetic bead (IMB) coated with ractopamine-single chain variable fragment (RAC-scFv). The intact RAC-scFv was constructed, and its key amino acid sites were analyzed. Finally, an icELISA based on immunomagnetic beads separation techniques (IMBs) of RAC from the porcine liver was established with an IC50 of 9.15 ng/mL and a linear range from 4.8 to 17.4 ng / mL. The IC50 of the prepared RAC-scFv was 8.7 ng/mL with a linear range of 3.8 to 19.6 ng/mL. The limit of detection (LOD) in the porcine liver of RAC was 4.57 μg/kg, and the recoveries ranged from 72.65 to 89.41% (CV < 15%).