Magneto-controlled potentiometric assay for E. coli based on cleavage of peptide by outer-membrane protease T

Abstract

Rapid, sensitive and reliable Escherichia coli (E. coli) detection and identification are critically important to protect public health. Here, we describe a magneto-controlled potentiometric assay for specific detection of E. coli cells by making use of the E. coli outer-membrane protease T (OmpT). OmpT is an endopeptidase that specifically cleaves peptide at dibasic sites. A rationally designed peptide serving as both OmpT substrate and potentiometric signal reporter was immobilized on magnetic beads. The rapid accumulation and extraction of peptide-functionalized magnetic beads on a polymeric membrane doped with an ion exchanger can be achieved using a magnetic force. The magnetic-field-assisted extraction of the peptide into the polymeric membrane ion-sensitive sensor, as confirmed by Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy, can lead to a rapid, stable and reproducible potential change. OmpT is capable of cleaving the positively charged peptide on the magnetic beads, thus resulting in charge density change. The change in charge density and subsequently the potential change can be readily detected and used for quantification of E. coli at levels down to 5.0 × 103 CFU mL−1. This work provides a versatile, rapid and reliable potentiometric method for E. coli detection.