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Legionellosis has been and continues to be a life-threatening disease worldwide, even in developed countries. Given the severity and unpredictability of Legionellosis outbreaks, developing a rapid, highly specific, and sensitive detection method is thus of great pertinence. In this paper, we demonstrate that sub-femtomole levels of 16s rRNA from pathogenic Legionella pneumophila can be timely and effectively detected using an appropriate designed capture, detector probes, and a QD SPRi signal amplification strategy. To achieve specific and sensitive detection, optimal hybridization conditions and parameters were implemented. Among these parameters, fragmentation of the 16s rRNA and further signal amplification by QDs were found to be the main parameters contributing to signal enhancement. The appropriate design of the detector probes also increased the sensitivity of the detection system, mainly due to secondary structure of 16s rRNA. The use of 16s rRNA from L. pneumophila allowed for the detection of metabolically active pathogens with high sensitivity. Detection of 16s rRNA in solutions as diluted as 1 pM at 450 μL (0.45 femtomole) was achieved in less than 3h, making our approach suitable for the direct, timely, and effective detection of L. pneumophila within man-made water systems., (Copyright © 2013 Elsevier B.V. All rights reserved.)

The use of dielectric spectroscopy to carry out real time observations of cells and to extract a wealth of information about their physiological properties has expanded in recent years. This popularity is due to the simple, easy to use, non-invasive and real time nature of dielectric spectroscopy. The ease of integrating dielectric spectroscopy with microfluidic devices has allowed the technology to further expand into biomedical research. Dielectric spectra are obtained by applying an electrical signal to cells, which is swept over a frequency range. This review covers the different methods of interpreting dielectric spectra and progress made in applications of impedance spectroscopy for cell observations. First, methods of obtaining specific electrical properties of cells (cell membrane capacitance and cytoplasm conductivity) are discussed. These electrical properties are obtained by fitting the dielectric spectra to different models and equations. Integrating models to reduce the effects of the electrical double layer are subsequently covered. Impedance platforms are then discussed including electrical cell substrate impedance sensing (ECIS). Categories of ECIS systems are divided into microelectrode arrays, interdigitated electrodes and those that allow differential ECIS measurements. Platforms that allow single cell and sub-single cell measurements are then discussed. Finally, applications of impedance spectroscopy in a range of cell observations are elaborated. These applications include observing cell differentiation, mitosis and the cell cycle and cytotoxicity/cell death. Future applications such as drug screening and in point of care applications are then covered., (Copyright © 2013. Published by Elsevier B.V.)

The advances in genomics and proteomics have unveiled an exhaustive catalogue of biomarkers that can potentially be used as diagnostic and prognostic indicators of genetic and infectious diseases. Current thrust in biosensor development is towards rapid, real-time, label-free and highly sensitive detection of the indicative biomarkers. While surface plasmon resonance imaging (SPRi) biosensors could potentially be the best suited candidate for biomarker-based diagnosis, important milestones need to be reached. Commercially available SPRi instrumentation is currently limited by the flow-cell technology to serial-sample processing and has limited sensitivity for the detection of markers present at low concentration. In this paper, we have implemented an approach to enhance sample handling and increase the sensitivity of the SPRi detection technique. We have developed a digital microfluidic platform with an integrated nanostructured biosensor interface that allows for rapid, ultra-low volume, sensitive, and automated on-chip SPRi detection of DNA hybridization reactions. Through the exploitation of electromagnetic properties of nanofabricated periodic gold nanoposts, SPRi signal was increased by 200% with the estimated limit of detection of 500 pM (90 attomoles). Using the versatile fluidic manipulation provided by the digital microfluidics, rapid and parallel target identification was achieved on multiple array elements within 1 min using 180 nL sample volume. By delivering multiple target analytes in individually addressable low volume droplets, without external pumps and fluidic interconnects, the overall assay time, cost and complexity was reduced. The proposed platform allows extreme versatility in the manipulation of precious low volume samples which makes this technology very suitable for diagnostic applications., (Copyright © 2010 Elsevier B.V. All rights reserved.)

In this report, nano-gratings with guided adsorption of biomolecules are investigated as new transducer elements or biointerfaces for surface plasmon resonance biosensor technologies. SPR biosensors are of particular interest due to the interaction between the electromagnetic fields and periodic nano-structures. In this article, sensitivity enhancement is demonstrated for a surface plasmon resonance interface, in a Kretschmann's configuration, featuring nano-gratings combined with nano-patterned immobilization of surface bioreceptors. The fabrication of this enhanced biointerface is demonstrated using a combination of metal lift-off and self-assembled monolayers. Rigorous coupled-wave analyses point to an increase in SPR angular response for the immobilization of surface bioreceptors onto areas of the nano-corrugated surface exhibiting high electromagnetic field intensity. Experimental measurements of the immobilization of anti-TNF-alpha antibody as a model bioreceptor using an imaging-SPR technique show a 3 times increase in angular resonance response from nano-grating surfaces with functionalized mesas compared to a planar surface or to a uniformly functionalized nano-grating surface. Furthermore, results also show an increased detection of TNF-alpha due to the increased accessibility to the adsorbed bioreceptors on the nano-gratings.

This work reports on a dynamically configurable micro-array surface plasmon resonance biochip platform. The platform comprises a digital electrowetting-on-dielectric (EWOD) microfluidic device tailored to surface plasmon resonance imaging (SPRi). We demonstrate its application for simultaneous immobilization of different DNA probes at the designated detection sites on-chip from sub-microL volume solutions in combination with multichannel label-free real-time detection of subsequent hybridization reactions. Successful on-chip DNA probe dilution and immobilization is also demonstrated using SPRi hybridization detection. Furthermore, active control of the immobilized probe density and orientation is achieved under an applied potential using the electric interface of the EWOD device. For low probe densities, under negative applied potential, the DNA hybridization efficiency is enhanced compared to passive probe immobilization, yielding a two-fold SPR signal increase within only 8min of hybridization. EWOD microfluidic platform coupled with SPRi promises to dramatically increase the speed of detection and quantification of biomolecular interactions while reducing reagent consumption. The proposed system would enable the development of high-throughput, rapid and ultrasensitive detection of biomolecules beyond DNA microarray applications.

Surfaces featuring nano-structures and biochemical patterns are increasingly developed as novel and superior substrates for biosensors and assays. Metallic periodic nano-structures have been studied for their unique optical properties and in particular their ability to support surface plasmon waves. Here we present a new nano-structuring approach based on gentle metal lift-off process coupled with self-assembled surface chemistry for the fabrication of a zeroth-order 400nm period metallic grating with differentiated surface chemistries on the mesas and troughs. The approach, using terminated self-assembled monolayers, creates versatile functionalized substrates allowing the precise deposition of complex biomolecular structures. We use this technique to perform the guided deposition of a three-dimensional polyelectrolyte multilayer structure and the patterned adsorption of quantum dots. Finally, we demonstrate that scanning near-field optical microscopy, used in conjuncture with atomic force microscopy and scanning electron microscopy, is an ideal tool for the characterization of this nano-structured surface as it provides a complete chemical, topographical and optical image of the surface. This ability to pattern and locally measure the surface properties is likely to have an important impact on the design of novel and optimized biointerfaces and transducers for biosensors.

The use of surface plasmon resonance (SPR) biosensors is increasingly popular in fundamental biological studies, health science research, drug discovery, clinical diagnosis, and environmental and agricultural monitoring. SPR allows for the qualitative and quantitative measurements of biomolecular interactions in real-time without requiring a labeling procedure. Today, the development of SPR is geared toward the design of compact, low-cost, and sensitive biosensors. Rapid advances in micro-fabrication technology have made available integratable opto-electronic components suitable for SPR. This review paper focuses on the progress made over the past 4 years toward this integration. Readers will find the descriptions of novel SPR optical approaches and materials. Nano-technology is also increasingly used in the design of biologically optimized and optically enhanced surfaces for SPR. Much of this work is leading to the integration of sensitive SPR to lab-on-a-chip platforms.