The Development of A New, Rapid, Amperometric Immunosensor for the Detection of Low Concentrations of Bacteria Part I: Design of the Detection System and Applications

Abstract

A portable, flow-through amperometric immunoassay system utilizing a newly designed immunosensor has been developed for the detection of bacteria. A sandwich scheme of immunoassay involving two stages of immunoreaction was employed using Escherichia coli as a model. E. coli antibodies were immobilized on Toray carbon paper to create a disposable immunofiltration membrane. Detection of the bacteria was achieved through the use of horseradish peroxidase-labeled antibodies, which, when coupled with a substrate of hydrogen peroxide and sodium iodide, yielded an electro-reduction, on the order of microamps, which was measured amperometrically. The immunosensor is highly sensitive and selective through the use of the disposable immunofiltration membranes that also function as immunoelectrodes, which enhance the rate of the immunoreaction due to the high surface to volume ratio of the solid to liquid phases. Potential commercialization through easier sample preparation and higher reproducibility of this system was a major objective, so the design highlights simplicity along with functionality. Preliminary data shows concentrations as low as 50 cell/mL can be detected in 22 min. Bacteria can be detected but not quantified over the desired working range of 50-1000 cell/mL. Optimization of the system parameters must be completed in order to ensure correct functionality of the sensor over the desired range of detection.