The effect of varying the peptide linker length in a single chain variable fragment antibody against wogonin glucuronide

•Three single-chain variable fragments to wogonin glucuronide (Wgn) were prepared.•The scFvs having different linker lengths showed the similar characteristics.•The indirect competitive ELISA using the scFv was developed for detection of Wgn.•The contents of Wgn in Scutellariae Radixes were analyzable by the icELISA.•The scFv is low-cost, effective, and applicable for improvement of antibodies.

Peptide linkers of three different lengths were constructed to join the variable regions of the heavy chain (VH) and the light chain (VL) in a single-chain variable fragment antibody (scFv) specific for wogonin glucuronide (Wgn) that has the structure VH-(GGGGS)n-VL (n = 3, 5, or 7). The scFv antibodies, which were expressed in Escherichia coli, were derived from an anti-Wgn monoclonal antibody (315A). An indirect competitive enzyme-linked immunosorbent assay (icELISA) was used to evaluate their reactivity and sensitivity, which is also used for quantitative analysis of Wgn. Our results, showed that the reactivity and specificity of the three different scFvs were, in fact, similar. Subsequently, the scFv having a VH-(GGGGS)3-VL linker which was slightly better that other two scFvs against Wgn, was applied to indirect competitive ELISA (icELISA) to analyze Scutellariae Radix (S. Radix). The utility of the icELISA was demonstrated for quality control and analysis of S. Radix in this report.