Simultaneous Electrochemical Detection of Multiple Bacterial Species Using Metal-Organic Nanohybrids.

Organic metallic nanohybrids (NHs), in which many small metal nanoparticles are encapsulated within a conductive polymer matrix, are useful as sensitive electrochemical labels because the constituents produce characteristic oxidation current responses. Gold NHs, consisting of gold nanoparticles and poly(m-toluidine), and copper NHs, consisting of copper nanoparticles and polyaniline, did not interfere with each other in terms of the electrochemical signals obtained on the same electrode. Antibodies were introduced into these NHs to function as electrochemical labels for targeting specific bacteria. Electrochemical measurements using screen-printed electrodes dry-fixed with NH-labeled bacterial cells enabled the estimation of bacterial species and number within minutes, based on the distinct current response of the labels. Our proposed method achieved simultaneous detection of enterohemorrhagic Escherichia coli and Staphylococcus aureus in a real sample. These NHs will be powerful tools as electrochemical labels and are expected to be useful for rapid testing in food and drug-related manufacturing sites.

Simultaneous Optical Detection of Multiple Bacterial Species Using Nanometer-Scaled Metal-Organic Hybrids.

This paper describes a simple strategy to identify bacteria using the optical properties of the nanohybrid structures (NHs) of polymer-coated metal nanoparticles (NPs). NHs, in which many small NPs are encapsulated in polyaniline particles, are useful optical labels because they produce strong scattered light. The light-scattering characteristics of NHs are strongly dependent on the constituent metal elements of NPs. Gold NHs (AuNHs), silver NHs (AgNHs), and copper NHs (CuNHs) produce white, reddish, and bluish scattered light, respectively. Moreover, unlike NPs, the color of the scattered light does not change even when NHs are aggregated. Introducing an antibody into NHs induces antigen-specific binding to cells, enabling the identification of bacteria based on light scattering. Multiple bacterial species adsorbed on the slide can be identified within a single field of view under a dark field microscope based on the color of the scattered light. Therefore, it is a useful development for safety risk assessments at manufacturing sites, such as those for foods, beverages, and drugs, and environmental surveys that require rapid detection of multiple bacteria.

Development of highly sensitive optical nanoantenna for bacterial detection.

Gold nanoparticles (AuNPs) are chemically stable and serve as excellent labels because their characteristic red coloration based on the localized surface plasmon resonance (LSPR) does not fade. However, it is necessary to control the structure of AuNPs to use them as labels for various analyses, because their optical properties depend strongly on their size, shape, and state of aggregation. In this study, we developed gold nanostructures (AuNSs) by encapsulating many small AuNPs within a polymer for scattering light-based bacterial detection. The AuNSs consisting of many small nanoparticles provided stronger scattered light intensity than a single AuNP of the same particle size. We found that the aggregation of the AuNSs enhanced the scattering light intensity, depending strongly on their aggregation states, and did not affect the wavelength of the scattering light observed under a dark-field microscope. By specifically binding the antibody-introduced AuNSs to the antigen on the bacterial surface, it was possible to label the target bacteria and detect them based on their light scattering characteristics. In addition, to improve the accuracy of the selective identification of the cells of interest, labels based on scattered light should ideally have a fixed wavelength of scattered light with high intensity. From these perspectives, we developed a method of constructing an optical antenna on the surface of target bacterial cells using antibody-introduced NSs.

Quantification of Enterohemorrhagic Escherichia coli via Optical Nanoantenna and Temperature-responsive Artificial Antibodies.

Enterohemorrhagic Escherichia coli are a dangerous bacterium known to be harmful to the human body, with some infections even resulting in death. Given this danger, food factories are required to perform a quick bacterial test to confirm the absence of this pathogen prior to shipping. We have developed a novel molecular imprinting polymer (MIP) particle that has encapsulated gold nanoparticles (AuNPs) and which can function as both a receptor and an optical signal transmitter in biological systems. This MIP particle is artificially synthesized and can be engineered to specifically recognize and capture antigens on the bacterial cell membrane. In addition, MIP particles containing AuNPs generate strong scattered light signals, and binding of the MIP particles improves the optical intensity of the target bacterial cells. This enables clear visualization under a dark-field microscope and quantification of the target bacteria using the scattering light intensity. Here we describe the successful quantification of Escherichia coli O157 cells in real meat samples using this technology in conjunction with a simple labelling step.