Isolation and characterization of single-chain variable fragment (scFv) antibody against polymeric immunoglobulin receptor, a potential diagnostic biomarker for Opisthorchis viverrini-related cholangiocarcinoma

The Polymeric Immunoglobulin Receptor (PIGR) has been identified as a promising candidate for a diagnostic biomarker in Opisthorchis viverrini-related cholangiocarcinoma (CCA).Developing a single-chain variable fragment (scFv) antibody targeting PIGR could significantly advance diagnostic methodologies in CCA.Antibody phage display technology was used to screen for scFv antibodies specific to PIGR, employing the PIGR315-354 peptide as an antigen.Bio-panning was conducted using the Yamo I library.Positive phage clones were identified through phage enzyme-linked immunosorbent assay (ELISA) (indirect ELISA).The specificity of expressed scFv in Escherichia coli DH5 was subsequently confirmed by ELISA and dot-blot analysis.The DNA sequences of positive phage clones and their deduced amino acid sequences were analyzed.Furthermore, the scFv interaction with the PIGR peptide was predicted.The isolation of the PIGR-specific phage clone E4 was achieved.ELISA assays revealed that both the E4 bacteriophage clone and the expressed E4 scFv specifically bound to PIGR315-354 in comparison to BSA, PIGR225-268, and the PIK3CB2-43 peptide.Dot-blot analysis further confirmed the specific affinity of E4 towards PIGR315-354.The deduced amino acid sequences of the E4 bacteriophage clone indicated a full-length scFv antibody.Protein structure prediction clearly indicated the presence of the V H and V L domains, supporting the specificity of the isolated E4 scFv antibody for the PIGR315-354 peptide.The E4 scFv antibody, derived from the Yamo I library, exhibits specificity for PIGR315-354.This scFv antibody holds significant potential for developing diagnostic methods for CCA, presenting promising prospects for future diagnostic approaches. KEYWORDS: polymeric immunoglobulin receptor (PIGR),