Development of an immunosensor using oriented immobilized anti-OmpW for sensitive detection of Vibrio cholerae by...

The first SPR sensor for detection of bacteria was reported in 1998 with high detection limit as much as 10(7)cfu/mL. Since then, a lot of effort has been made to lower detection limit and increase sensitivity of detection mainly by using of different assay formats, immobilization strategies, suitable antibodies, minimizing non-specific adsorption and improving the quality of SPR devices. The aim of this paper is to introduce the potential of an antibody against recombinant outer membrane protein (anti-OmpW) in sensitive detection of Vibrio cholerae by developing an immunosensor based on SPR and compare the sensitivity of this method with former report for detection of V. cholerae published in 2006. Recombinant OmpW antigen (a bacterial outer-membrane protein) of V. cholerae was expressed and purified and raising of polyclonal rabbit anti-OmpW was done. Protein G was covalently immobilized on 11-MUA SAM via amine coupling and bioaffinity-based oriented immobilization of anti-OmpW was done on protein G layer. The results showed high affinity interaction between OmpW and anti-OmpW (KD=2.4×10(-9)M) and the detection limit of fabricated immunosensor was 43 cells/mL. The apparent reasons for achieving this low LOD are discussed.