Detection Of DNA Hybridization Events Research Articles

Effects of Amine Linkers with Different Carbon Chain Lengths at Guanine-Rich Polynucleotides on Chemical Interface Damping in Single Gold Nanorods.

DNA-functionalized gold nanoparticles (AuNPs) are used for various bioapplications, such as biosensor development and drug delivery. Nevertheless, no study has reported the effect of polynucleotide chains on chemical interface damping (CID), the most recently proposed plasmon damping pathway in single AuNPs. In this study, we conducted total internal reflection scattering measurements of gold nanorods (AuNRs) to reveal the CID effect induced by amine (NH2)-linked polynucleotides (or DNA) with guanine-rich sequences through the interaction between nitrogen and Au surfaces. Additionally, we elucidated the effect of a linear hydrocarbon chain length between NH2 and DNA (NH2-Cn-DNA, n = 6, 12, 18, 24) on spectral changes in single AuNRs. The localized surface plasmon resonance (LSPR) linewidth increased with an increasing number of linear carbon, from 6 to 24, due to the increase in van der Waals forces. Second, the effect of the direction (5' or 3' ends) of DNA attachment to the AuNR surfaces on LSPR spectral changes was investigated, and there was no significant difference in LSPR wavelength and full linewidth at half-maximum shifts caused by the DNA attachment directions (5' or 3' ends). Third, guanine-rich DNA can fold into four-stranded secondary structures called G-quadruplexes (GQs). We demonstrated the effect of linear carbon chain length, between NH2 and GQs, on CID in single AuNRs. Lastly, a label-free detection of DNA hybridization events on single AuNRs was demonstrated for sensing applications. Thus, we provide an insight into the effect of amine-functionalized guanine-rich DNA with different carbon chains on LSPR spectral changes, including CID in single AuNRs.

Detection of DNA hybridization using graphene-coated black phosphorus surface plasmon resonance sensor

In this paper, graphene-coated black phosphorus at the metal surface for the detection of DNA hybridization event is numerically demonstrated. The strategy consists of placing the sensing medium on top of black phosphorus–graphene-coated SPR which interfaces with phosphate-buffered saline solution carrying single-stranded DNA. Upon hybridization with its complementary DNA, desorption of the nanostructures takes place and thus enables the sensitive detection of the DNA hybridization event. The proposed sensor exhibits a sensitivity (125 ο/RIU), detection accuracy (0.95) and quality factor (13.62 RIU−1) for complementary DNA. In comparison with other reported papers, our suggested sensor provides much better performance. Thus, this label-free DNA detection platform should spur off new interest towards the use of black phosphorus–graphene-coated SPR interfaces.

Microfluidic Arrayed Lab-On-A-Chip for Electrochemical Capacitive Detection of DNA Hybridization Events.

A microfluidic electrochemical lab-on-a-chip (LOC) device for DNA hybridization detection has been developed. The device comprises a 3×3 array of microelectrodes integrated with a dual layer microfluidic valved manipulation system that provides controlled and automated capabilities for high throughput analysis of microliter volume samples. The surface of the microelectrodes is functionalized with single-stranded DNA (ssDNA) probes which enable specific detection of complementary ssDNA targets. These targets are detected by a capacitive technique which measures dielectric variation at the microelectrode-electrolyte interface due to DNA hybridization events. A quantitative analysis of the hybridization events is carried out based on a sensing modeling that includes detailed analysis of energy storage and dissipation components. By calculating these components during hybridization events the device is able to demonstrate specific and dose response sensing characteristics. The developed microfluidic LOC for DNA hybridization detection offers a technology for real-time and label-free assessment of genetic markers outside of laboratory settings, such as at the point-of-care or in-field environmental monitoring.

Label-free DNA biosensor based on resistance change of platinum nanoparticles assemblies

A novel nanoparticle based biosensor for the fast and simple detection of DNA hybridization events is presented. The sensor utilizes hybridized DNA's charge transport properties, combining them with metallic nanoparticle networks that act as nano-gapped electrodes. The DNA hybridization events can be detected by a significant reduction in the sensor's resistance due to the conductive bridging offered by hybridized DNA. By modifying the nanoparticle surface coverage, which can be controlled experimentally being a function of deposition time, and the structural properties of the electrodes, an optimized biosensor for the in situ detection of DNA hybridization events is ultimately fabricated. The fabricated biosensor exhibits a wide response range, covering four orders of magnitude, a limit of detection of 1nM and can detect a single base pair mismatch between probe and complementary DNA.

Highly sensitive detection of DNA hybridization on commercialized graphene-coated surface plasmon resonance interfaces.

Strategies employed to interface biomolecules with nanomaterials have considerably advanced in recent years and found practical applications in many different research fields. The construction of nucleic acid modified interfaces together with the label-free detection of hybridization events has been one of the major research focuses in science and technology. In this paper, we demonstrate the high interest of graphene-on-metal surface plasmon resonance (SPR) interfaces for the detection of DNA hybridization events in the attomolar concentration range. The strategy consists on the noncovalent functionalization of graphene-coated SPR interfaces with gold nanostars carrying single-stranded DNA (ssDNA). Upon hybridization with its complementary DNA, desorption of the nanostructures takes place and thus enables the sensitive detection of the DNA hybridization event. The DNA sensor exhibits a detection limit of ≈500 aM for complementary DNA with a linear dynamic range up to 10(-8) M. This label-free DNA detection platform should spur off new interest toward the use of commercially available graphene-coated SPR interfaces.

Localized surface plasmon resonance (LSPR) biosensing using gold nanotriangles: detection of DNA hybridization events at room temperature.

We present a proof-of-concept of the application of gold nanotriangles in sequence specific DNA detection, using localized surface plasmon resonance (LSPR) spectroscopy and dark-field optical microscopy. The sensing platform comprises gold nanotriangles immobilized on a glass chip and oligonucleotides as probes. Probe formation and testing complementary and non-complementary targets followed common chip technology protocols. Gold nanotriangles showed a remarkable sensitivity of 468 nm per RIU and allowed detection of 20-mer targets. When the target sequence was part of a 50-mer synthetic DNA oligonucleotide, LSPR shifts as high as 35 nm were observed. Conversely, when the target was present in PCR products of ca. 350 bp, obtained from clinical samples, LSPR shifts larger than 20 nm were observed. Moreover, LSPR shifts were less than ±1 nm for the respective non-complementary targets. These results with gold nanotriangles as sensors are a notable improvement to the LSPR shifts of less than 5 nm usually obtained for spherical gold nanoparticles of comparable sizes. Optimal conditions for the detection of synthetic and PCR product targets using gold nanotriangles and oligonucleotide probes were achieved with low percentages of intercalating thioalkanes; target hybridization at room temperature, 3 hours of incubation, and 2× SSC buffer stringency conditions.

Photoinitiator Nucleotide for Quantifying Nucleic Acid Hybridization

This first report of a photoinitiator-nucleotide conjugate demonstrates a novel approach for sensitive, rapid and visual detection of DNA hybridization events. This approach holds potential for various DNA labeling schemes and for applications benefiting from selective DNA-based polymerization initiators. Here, we demonstrate covalent, enzymatic incorporation of an eosin-photoinitiator 2′-deoxyuridine-5′-triphosphate (EITC-dUTP) conjugate into surface-immobilized DNA hybrids. Subsequent radical chain photoinitiation from these sites using an acrylamide/bis-acrylamide formulation yields a dynamic detection range between 500pM and 50nM of DNA target. Increasing EITC-nucleotide surface densities leads to an increase in surface-based polymer film heights until achieving a film height plateau of 280nm ±20nm at 610 ±70 EITC-nucleotides/μm2. Film heights of 10–20 nm were obtained from eosin surface densities of approximately 20 EITC-nucleotides/μm2 while below the detection limit of ~10 EITC-nucleotides/μm2, no detectable films were formed. This unique threshold behavior is utilized for instrument-free, visual quantification of target DNA concentration ranges.

Fluorescence Lifetime Imaging of Quantum Dot Labeled DNA Microarrays

Quantum dot (QD) labeling combined with fluorescence lifetime imaging microscopy is proposed as a powerful transduction technique for the detection of DNA hybridization events. Fluorescence lifetime analysis of DNA microarray spots of hybridized QD labeled target indicated a characteristic lifetime value of 18.8 ns, compared to 13.3 ns obtained for spots of free QD solution, revealing that QD labels are sensitive to the spot microenvironment. Additionally, time gated detection was shown to improve the microarray image contrast ratio by 1.8, achieving femtomolar target sensitivity. Finally, lifetime multiplexing based on Qdot525 and Alexa430 was demonstrated using a single excitation-detection readout channel.

Label-free DNA detection based on a novel nanostructured conducting poly(indole-6-carboxylic acid) films

A simple and label-free electrochemical sensor for detection of DNA hybridization event was obtained based on a novel nanostructured conducting polymer, poly(indole-6-carboxylic acid). This precursor polymer with high electroactivity can be easily prepared by direct electrochemical polymerization of indole-6-carboxylic acid in acetonitrile containing tetrabutylammonium tetrafluoroborate. An 18-mer amino-substituted oligonucleotide (ODN) probe was covalently grafted onto the polymer in a one-step procedure. The performance of this label-free electrochemical DNA sensor was investigated by using cyclic voltammetry (CV) as a readout method. This simple sensor can discriminate one-base mismatched and non-complementary ODN sequences and can be regenerated after a dehybridization step. The dynamic determination range for complementary target ODN was from 3.5 × 10 −10 mol L −1 to 2.0 × 10 −8 mol L −1 and the corresponding detection limit was 5.79 pmol L −1.

Spermine-Induced Hybridization and Charge Inversion at the Diffuse Layer of a DNA-FET

Detection of DNA hybridization events by using field-effect transducers is limited by the electrolyte content of the medium. So far, DNA was thought to hybridize only in solutions with concentrated electrolytes. In these media, the interface between the transducer gate and the solution is reduced to a thin layer in close contact with the surface, and DNA is poorly detected. In the present work, this limitation is overcome by using spermine as screening polycation. Hybridization assays with polycation concentrations as low as 10 microM are reported. This ensures that hybridization takes place at the diffuse layer of the interface. The reported results suggest a charge-inversion mechanism induced by spermine. A target sequence is real-time label-free detected in the range from 10 to 500 nM.

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Nanoporous niobium oxide for label-free detection of DNA hybridization events

We found that DNA probes can be immobilized on anodically prepared porous niobium oxide without a chemical modification of both the DNA probes and the substrate. By using the porous niobium oxide with a positive surface charge, DNA hybridization events are detected on the basis of the blue-shift of a maximum absorption peak in UV–vis-NIR spectroscopy. The blue-shift is ascribed to the change of surface charge upon single- or double-stranded DNA. The method does not require a label and shows high sensitivity with the detection limit of the concentration of 1 nM.

Dendron Arrays for the Force-Based Detection of DNA Hybridization Events

Single-molecule force measurement methods have attracted increasing interest over recent years for the development of novel approaches for biomolecular screening. However, many of these developments are currently hindered by the available biomolecule surface attachment methods, in that it is still not trivial to create surfaces and devices with highly defined surface functionality and/or uniformity. Here we offer a new approach to address such issues based on the formation of dendron arrays. Through the measurement of forces between dendron surfaces functionalized with complementary DNA oligonucleotides, we observed several unique properties of the surfaces modified via this approach. The capability to record attractive or "jump-in" forces associated with molecular binding events is one of them. Additionally, these events occur in greater than 80% of measurements, and the forces are dependent on the number of complementary DNA bases of the associating strands while being insensitive to the measurement rate. Combined with a narrow distribution of both attractive forces and unbinding forces we suggest such functionalized surfaces offer a significant advance for fast and accurate force-based studies of oligonucleotide hybridization.

Electrochemical studies of (−)-epigallocatechin gallate and its interaction with DNA

In this paper, an electrochemical investigation of (-)-epigallocatechin gallate (EGCG) and its interaction with DNA is presented. Via an electrochemical approach assisted by ultraviolet-visible (UV-Vis) spectroscopy, we propose that EGCG can intercalate into DNA strands forming a nonelectroactive complex, which results in the decrease of the anodic peak current of EGCG. Meanwhile, an electrochemical study with the DNA-Cu(II)-EGCG system shows that damage to DNA can be recognized electrochemically via the increase in the anodic peak current resulting from the oxidation of guanine and adenine bases. The damage can also be recognized spectrophotometrically via an increase in the 260 nm absorption band. In addition, it was found that EGCG is able to discriminate dsDNA from ssDNA, making a potential electrochemical indicator for the detection of DNA hybridization events. A rapid and convenient method of detecting EGCG was also developed in this work.

Electronic detection of single-base mismatches in DNA with ferrocene-modified probes.

Genotyping and gene-expression monitoring is critical to the study of the association between genetics and drug response (pharmacogenomics) and the association of sequence variation with heritable phenotypes. Recently, we developed an entirely electronic method for the detection of DNA hybridization events by the site-specific incorporation of ferrocenyl derivatives into DNA oligonucleotides. To perform rapid and accurate point mutation detection employing this methodology, two types of metal-containing signaling probes with varying redox potentials are required. In this report we describe a new ferrocene-containing phosphoramidite 9 that provides a range of detectable redox potentials. Using automated DNA/RNA synthesis techniques the two ferrocenyl complexes were inserted at various positions along oligonucleotide probes. Thermal stability analysis of these metal-containing DNA oligonucleotides indicates that incorporation of 9 resulted in no destabilization of the duplex. A mixture of oligonucleotides containing compounds 9 and I was analyzed by alternating current voltammetry (ACV) monitored at the 1st harmonic. The data demonstrate that the two ferrocenyl oligonucleotide derivatives can be distinguished electrochemically. A CMS-DNA array was prepared on an array of gold electrodes on a printed circuit board substrate with a self-assembled mixed monolayer, coupled to an electronic detection system. Experiments for the detection of a single-base match utilizing two signaling probes were carried out. The results demonstrate that rapid and accurate detection of a single-base mismatch can be achieved by using these dual-signaling probes on CMS-DNA chips.