A tetravalent single-chain variable fragment antibody for the detection of staphylococcal enterotoxin A
Weifeng CHEN, Zhiwei LI, Xingxing DONG, Xiaohong WANG
School of Food and Bioengineering, Henan University of Animal Husbandry and Economy, Zhengzhou 450046, China; Key Laboratory of Environment Correlative Dietology, College of Food Science and Technology, Huazhong Agricultural University, Wuhan 430070, China
wxh@mail.hzau.edu.cn
Abstract: Staphylococcal enterotoxin A (SEA) synthesized by Staphylococcus aureus is a foodborne and heat-stable toxin, which is a great threat to human health (Pexaraet al., 2010). Highly sensitive antibodies are a key factor in the immunological detection of SEA, which is one of the most effective ways to detect SEA because of its accuracy, agility, and efficiency (Nouri et al., 2018). In this study, we constructed a tetravalent anti-SEA antibody gene by linking the tetramerization domain of human p53 to the C-terminus of the anti-SEA single-chain variable fragment (scFv), which was then transformed into Escherichia coli BL21 (DE3) for the production of a SEA-specific tetravalent antibody. Successful expression of the tetravalent antibody was determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and western blot. An indirect non-competitive enzyme-linked immunosorbent assay (ELISA) revealed that the tetravalent antibody exhibited SEA-specific binding activity. A sandwich ELISA demonstrated that compared to the scFv monomer, the tetravalent antibody was more sensitive in detecting SEA. Molecular docking analysis revealed that the SEA interacted with the scFv mainly on the opposite side of the residue linked to p53. Thus, this study indicated that genetically engineered tetramerization is a potential way to improve the sensitivity of SEA-specific scFv.