Electrochemical Immunosensor Using Nanoparticle-Based Signal Enhancement for <italic>Escherichia Coli</italic> O157:H7 Detection

Abstract

An electrochemical immunosensor based on the assembly of 3 nanoparticles for the rapid detection of Escherichia coli O157:H7 has been developed. The biosensor assay consists of magnetic separation using antibody-functionalized magnetic nanoparticles (MNPs) and electrochemical reporters using gold nanoparticle (AuNP)-conjugated lead sulfide (PbS) nanoparticles via oligonucleotide linkage. The gold nanoparticles (AuNPs) were also functionalized with polyclonal anti-E. coli O157:H7 antibodies in order to bind the target bacterial cells which were captured and separated from the sample by antibodyfunctionalized MNPs. Because each AuNP was linked to multiple PbS nanoparticles, each binding event to the target resulted in substantial amplification. The signal of PbS was measured on screen-printed carbon electrode (SPCE) by square wave anodic stripping voltammetry (SWASV). Results show that the biosensor could detect E. coli O157:H7 in the range of 10 1 to 10 6 colony forming units per milliliter (cfu/ml) with a signal-to-noise ratio ranging from 2.77 to 4.31. With sample preparation being minimized, results were obtained in 1 h from sample processing to final readout. Due to its high sensitivity, this tri-nano immunosensor has great potential applications in public health, biodefense, and food/water safety monitoring.

Culture‐Independent Rapid Detection Methods for Bacterial Pathogens and Toxins in Food Matrices

“…While many chemical methods and physicochemical methods have been investigated in foodborne separation and concentration (Stevens and Jaykus ), methods based on biological interactions such as immunomagnetic separation (IMS) has become one major approach for separating foodborne bacteria followed by various detection methods (Uyttendaele and others ; Gehring and others ; Ravindranath and others ; Zhao and others ; Yang and others ; Wang and others ). Besides antibody–antigen interaction, ligands such as aptamers (Joshi and others ; Wu and others 2014a), bacteriophage proteins (Poshtiban and others ), and other binding molecules (El‐Boubbou and others ) have been attached to magnetic beads for bacteria or bacterial nucleic acid (Zhang and others ; Li and others 2011b) separation in a similar way as IMS.…”

“…Besides antibody–antigen interaction, ligands such as aptamers (Joshi and others ; Wu and others 2014a), bacteriophage proteins (Poshtiban and others ), and other binding molecules (El‐Boubbou and others ) have been attached to magnetic beads for bacteria or bacterial nucleic acid (Zhang and others ; Li and others 2011b) separation in a similar way as IMS. The magnetic‐based separations have the advantages of specific removal of the target from food particles, inhibitors, and other organisms using rapid and easy manipulation with a common separation time within 1 h (Joshi and others ; Wang and others ; Poshtiban and others ; Cloutier and others ; Wang and others ), good recovery of the target (Uyttendaele and others ; El‐Boubbou and others ; Ravindranath and others ; Yang and others ; Wang and others ), no dilution of the sample or loss of carrier during washing, and possible automation and scale‐up separation (Horák and others ; Lau and others ; Chen and others ). The efficiency and selectivity of magnetic separation greatly depends on the ligands, as sometimes it is hard to obtain a ligand with high affinity and specificity to the target.…”

Screen-Printed Electrodes Modified with “Green” Metals for Electrochemical Stripping Analysis of Toxic Elements

“…The AuNPs were also functionalized with polyclonal anti- E. coli O157:H7 in order to bind the target bacterial cells, which were captured and separated from the sample by antibody-functionalized magnetic nanoparticles. The signal of PbS was measured witha Bi-coated SPE by SWASV [ 128 ]. A similar procedure was used to detect Bacillus anthracis [ 129 ] and this work was extended to duplex detection of the protective antigen A (pagA) gene of Bacillus anthracis and the insertion element (Iel) gene of Salmonella enteritidis [ 130 ].…”

A Reusable Column Method Using Glycopolymer‐Functionalized Resins for Capture–Detection of Proteins and Escherichia coli

“…Major approaches for separating food‐borne bacteria and their concentration include those based on biological interactions such as immunomagnetic separation (IMS), [ 26–31 ] antigen–antibody interaction, ligands such as aptamers, [ 32,33 ] bacteriophages, [ 34 ] and other binding molecules. [ 33 ] Magnetic‐bead‐based separation for bacteria has several advantages, like specific removal of target from food particles, [ 34–36 ] no dilution of sample or carrier during washing, and possible automation and scale‐up separation.…”