Generation and characterisation of a novel recombinant scFv antibody specific for Campylobacter jejuni

Campylobacter jejuni is a leading cause of foodborne illness worldwide, mainly due to consumption and handling of contaminated raw chicken. Rapid detection methods for C. jejuni are vital for monitoring contamination levels in chicken products and reducing human Campylobacteriosis cases. The ‘gold standard’ culture-based method of Campylobacter detection takes 3-5 days and is too slow to permit effective intervention. Immuno-based methods are faster, but usually necessitate use of animals or hybridoma technology to produce antibodies; making them difficult and expensive to produce. Here we report the generation and characterization of recombinant single-chain variable fragment (scFv) antibodies specific for C. jejuni cells, and evaluation of one scFv antibody for an immunomagnetic separation-quantitative PCR (IMS-qPCR) method to rapidly, sensitively and specifically detect low numbers of C. jejuni. An scFv antibody phage-display library was constructed using spleen mRNA derived from a rabbit immunised with gamma-irradiated C. jejuni cells. This library was screened by surface biopanning against C. jejuni whole cells. Enriched clones were analysed by enzyme-linked immunosorbent assay (ELISA). Two scFv antibodies that strongly and specifically recognised C. jejuni cells were expressed in Escherichia coli. Western blot analysis showed that one antibody, scFv80, was expressed as a soluble protein and retained its strong binding affinity and specificity for C. jejuni cells. This recombinant monoclonal scFv antibody was purified and used to covalently coat paramagnetic beads to be used for IMS-qPCR. The IMS-qPCR method was able to specifically and sensitively detect C. jejuni in mixed cultures within 3 h.