Enzyme shadowing: using antibody–enzyme dually-labeled magnetic particles for fast bacterial detection

Abstract

Using magnetic particles and immunoseparation for target recovery and detection has been reported to improve the performance and detection limits of traditional analytical methods. For example, magnetic immunocapture can be coupled to detection in a sandwich format using an antibody (Ab) labeled with a reporter molecule or enzyme. In this work we demonstrate that simultaneous incorporation of capture and reporter biocomponents onto the sensing surface is possible and provides assays that are extremely fast and easy to carry out. As a proof of concept, we have produced dually-labeled magnetic particles, simultaneously functionalized with antibody and reporter enzyme. Subsequent capture of Escherichia coli generates a shadowing effect on the particle surface and interferes with the activity of a number of enzyme units. The decrease in signal recorded is proportional to the bacterial concentration and is specific for the target microorganism, with a detection limit of 10³–10⁴cell mL⁻¹ in an assay of about one hour.