Real-time ligation chain reaction for DNA quantification and identification on the FO-SPR.

Different assays have been developed in the past years to meet point-of-care diagnostic tests requirements for fast and sensitive quantification and identification of targets. In this paper, we developed the ligation chain reaction (LCR) assay on the Fiber Optic Surface Plasmon Resonance (FO-SPR) platform, which enabled simultaneous quantification and cycle-to-cycle identification of DNA during amplification. The newly developed assay incorporated FO-SPR DNA melting assay, previously developed by our group.

Emerging technologies for hybridization based single nucleotide polymorphism detection.

Detection of single nucleotide polymorphisms (SNPs) is a crucial challenge in the development of a novel generation of diagnostic tools. Accurate detection of SNPs can prove elusive, as the impact of a single variable nucleotide on the properties of a target sequence is limited, even if this sequence consists of only a few nucleotides. New, accurate and facile strategies for the detection of point mutations are therefore absolutely necessary for the increased adoption of point-of-care molecular diagnostics.

Affinity comparison of p3 and p8 peptide displaying bacteriophages using surface plasmon resonance.

Ever increasing demands in sensitivity and specificity of biosensors have recently established a trend toward the use of multivalent bioreceptors. This trend has also been introduced in the field of bacteriophage affinity peptides, where the entire phage is used as a receptor rather than the individual peptides. Although this approach is gaining in popularity due to the numerous advantages, binding kinetics of complete phage particles have never been studied in detail, notwithstanding being essential for the efficient design of such applications.

Digital microfluidics-enabled single-molecule detection by printing and sealing single magnetic beads in femtoliter droplets.

Digital microfluidics is introduced as a novel platform with unique advantages for performing single-molecule detection. We demonstrate how superparamagnetic beads, used for capturing single protein molecules, can be printed with unprecedentedly high loading efficiency and single bead resolution on an electrowetting-on-dielectric-based digital microfluidic chip by micropatterning the Teflon-AF surface of the device. By transporting droplets containing suspended superparamagnetic beads over a hydrophilic-in-hydrophobic micropatterned Teflon-AF surface, single beads are trapped inside the hydrophilic microwells due to their selective wettability and tailored dimensions.

Nucleic acids for ultra-sensitive protein detection.

Major advancements in molecular biology and clinical diagnostics cannot be brought about strictly through the use of genomics based methods. Improved methods for protein detection and proteomic screening are an absolute necessity to complement to wealth of information offered by novel, high-throughput sequencing technologies. Only then will it be possible to advance insights into clinical processes and to characterize the importance of specific protein biomarkers for disease detection or the realization of "personalized medicine".

Selection of aptamers against Ara h 1 protein for FO-SPR biosensing of peanut allergens in food matrices.

The rising prevalence to food allergies in the past two decades, together with the fact that the only existing therapy is avoidance of allergen-containing food next to the implementation of anti-allergic drugs, urges the need for improved performance of current assays to detect potential allergens in food products. Therein, the focus has been on aptamer-based biosensors in recent years. In this paper we report for the first time the selection of aptamers against one of the most important peanut allergens, Ara h 1.

Spherical nucleic acid enhanced FO-SPR DNA melting for detection of mutations in Legionella pneumophila.

A home-built fiber optic surface plasmon resonance platform (FO-SPR) was applied to directly screen PCR amplified DNA for mutations. The FO-SPR sensor was used for real-time monitoring of DNA duplex melting during high resolution temperature cycling. The signal of the DNA melting was enhanced by means of gold nanoparticle labels.

Multiplexed protein detection using an affinity aptamer amplification assay.

Affinity probe capillary electrophoresis (APCE) is potentially one of the most versatile technologies for protein diagnostics, offering an excellent balance between robustness, analysis speed and sensitivity. Combining the immunosensing and separating strength of capillary electrophoresis with the signal enhancement power of nucleic acid amplification, aptamers can further push the analytical limits of APCE to offer ultrasensitive, multiplexed detection of protein biomarkers, even when differences in electrophoretic mobility between the different aptamer-target complexes are limited. It is demonstrated how, through careful selection of experimental parameters, simultaneous detection of picomolar levels of three target proteins can be achieved even with aptamers that were initially selected under very different conditions and further taking into account that the aptamers need to be modified to allow successful PCR amplification.

Real-time monitoring of DNA hybridization and melting processes using a fiber optic sensor.

In this paper a fiber optic surface plasmon resonance (FO-SPR) sensor was used to analyze the melting process of DNA linked to silica nanoparticles. Real-time monitoring of a DNA melting process has rarely been studied using surface plasmon resonance (SPR), since most commercial SPR setups do not allow for dynamic and accurate temperature control above 50 °C. The FO-SPR sensor platform, with silica nanobead signal amplification, allows sensing inside a standard PCR thermocycler, which makes high resolution DNA melting curve analysis possible.

A simple double-bead sandwich assay for protein detection in serum using UV-vis spectroscopy.

In this study a double-bead sandwich assay, employing magnetic nanoparticles and gold nanoparticles is proposed. The magnetic nanoparticles allow specific capturing of the analyte in biological samples, while the optical properties of the gold nanoparticles provide the signal transduction. We demonstrated that a major improvement in the assay sensitivity was obtained by selecting an optimal gold nanoparticle size (60 nm).