Detection of Edwardsiella tarda by fluorometric or biosensor methods using a peptide ligand

Abstract

In this study, we identified a peptide ligand for Edwardsiella tarda from a phage peptide library and tested two approaches for sensitive detection of the bacteria with the peptide labeled with fluorescein or biotin. At first, the fluorescent peptide was proved to be advantageous in the fluorescence polarization (FP) assay because sensitivity of the assay is maximized when a fluorophore is linked to a small molecule. The FP assay using the fluorescent peptide enabled detection of E. tarda in a range from 5.2 × 10 3 to 2.1 × 10 5 cells. Second, we devised a new assay method using a quartz crystal microbalance (QCM) biosensor connected to a filter module. When a mixture of E. tarda and the biotinylated peptide was injected into the filter module, the E. tarda–peptide complex was separated from the unbound peptide by a filter and detected with a streptavidin-coated QCM sensor chip. On injection of samples containing the biotinylated peptide and E. tarda, concentration-dependent frequency change was observed in a range from 8 × 10 2 to 8 × 10 6 cells. The two approaches are expected to facilitate development of assay methods using other bacteria-binding peptides.