from Theron et al. in Nanotechnology in Water Treatment Applications

Metal nanoparticles, such as gold, silver and iron, constitute one of the most researched branches of nanotechnology due to their electronic, optical, catalytic and thermal properties. Among these, gold nanoparticles are intensively used in a variety of colorimetric and fluorescence biodetection assays. Specific focus has been directed at colloidal gold nanoparticles ranging from 3 to 100 nm in size, since they are rather stable and their properties can be easily tailored by chemical modification of their surfaces.

Gold nanoparticle-based probes have been used in the identification of pathogenic bacteria in a chip-based system. The assay consists of a capture DNA strand immobilized on a glass chip that recognizes the DNA of interest. A separate sequence on the captured target strand is then labelled with an oligonucleotide-functionalized gold nanoparticle probe. After catalytic reduction of silver onto the gold nanoparticle surfaces to amplify the target signal, the capture-strand/target/nanoparticle sandwich is visualized with a flatbed scanner. The target DNA was detected at concentrations of 50 fM, which represents a nearly 100-fold increase in sensitivity over traditional fluorescence-based assays. Gold nanoparticles have also been used in the detection of genomic DNA from Staphylococcus aureus, without enzymatic amplification, at concentrations of 33 fM. The assay depends upon the target-selective binding of gold nanoparticle probes to consecutive regions of a target DNA sequence in solution, followed by transfer to an evanescently illuminated glass microscope slide for light scattering measurements. Whereas the gold particle probes scatter orange light in the presence of target, the probes scatter green light if the target is absent, whilst the target concentration is quantified by measuring the intensity of scattered light. Multiplexed detection of different viruses and bacteria, using gold nanoparticles surface functionalized with a target-specific oligonucleotide and further encoded with a Raman-active dye molecule, has also been demonstrated. Whereas the complementary probe sequence imparts the specificity for the target sequence of interest, the presence of the target is confirmed by silver staining and the identity of the target is revealed by detecting the surface-enhanced Raman scattering (SERS) of the Raman dye near the nanoparticle surface. This assay demonstrated a sensitivity of 1 fM target concentration.

Despite its widespread use in nucleic acid hybridization assays, the use of gold nanoparticles for detection of bacterial pathogens in immunoassays has only recently been described. The assay comprises detection of organism-specific antigens with biotinylated polyclonal antibodies after which gold particles functionalized with a secondary antibiotin antibody are added, and the particles are then visualized under a dark-field stereomicroscope. The immunoassays were demonstrated to reliably detect Helicobacter pylori and E. coli O157:H7 antigens in quantities in the order of 10 ng, which provides a sensitivity of detection comparable to those of conventional dot blot assays. A two dot filter system has also been developed where colloidal gold nanoparticles (2 nm) were bounded onto anti-E.coli O157:H7 antibodies. These monoclonal antibodies were bounded onto a 0.2 microm nitrocellulose filtermembrane through which water was filtered. The same antibodies that captured the bacteria acted as detectors since the gold nanoparticles could be visualized under epifluorescence. This one-step detection method not only had a low detection rate (1 cfu/100 ml) and high specificity but would be inexpensive and easy implemented in the routine testing of water samples.

Tags: Microbial Detection | Pathogen Detection | Biodiagnostics | Biodetection Assays | Biomolecular Detection