Expression and characterization of single-chain variable fragment antibody against staphylococcal enterotoxin A in Escherichia coli

Abstract

The staphylococcal enterotoxins (SEs) are potent gastrointestinal exotoxins synthesized by Staphylococcus aureus, which is responsible for various diseases including septicemia, food poisoning, and toxic shock syndrome, as well as bovine mastitis. Among them, staphylococcal enterotoxin A (SEA) is one of the most commonly present serotypes in staphylococcal food poisoning cases. In this study, the stable hybridoma 3C12 producing anti-SEA monoclonal antibody was established with an equilibrium dissociation constant (K_D) of 1.48 × 10⁻⁸ mol·L⁻¹, its ScFv-coding genes were obtained and then the anti-SEA single chain variable fragment (ScFv) protein was expressed in Escherichia coli. Characterization of the expressed target ScFv protein was analyzed by sodium dodecyl sulfate – polyacrylamide gel electrophoresis, Western blot, and enzyme-linked immunosorbent assay. The results demonstrated that the recombinant anti-SEA ScFv protein retained a specific binding activity for SEA, and the K_D value of the soluble ScFv was about 3.75 × 10⁻⁷ mol·L⁻¹. The overall yield of bioactive anti-SEA ScFv in E. coli flask culture was more than 10 mg·L⁻¹.