Colorimetric adenosine aptasensor based on DNA cycling amplification and salt-induced aggregation of gold nanoparticles

An aptamer based assay is described for the colorimetric detection of adenosine. The presence of adenosine triggers the deformation of hairpin DNA oligonucleotide (HP1) containing adenosine aptamer and then hybridizes another unlabeled hairpin DNA oligonucleotide (HP2). This leads to the formation of a double strand with a blunt 3′ terminal. After exonuclease III (Exo III)-assisted degradation, the guanine-rich strand (GRS) is released from HP2. Hence, the adenosine-HP1 complex is released to the solution where it can hybridize another HP2 and initiate many cycles of the digestion reaction with the assistance of Exo III. This leads to the generation of a large number of GRS strands after multiple cycles. The GRS stabilize the red AuNPs against aggregation in the presence of potassium ions. If, however, GRS forms a G-quadruplex, it loses its ability to protect gold nanoparticles (AuNPs) from salt-induced AuNP aggregation. Therefore, the color of the solution changes from red to blue which can be visually observed. This colorimetric assay has a 0.13 nM detection limit and a wide linear range that extends from 5 nM to 1 μM.

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Graphical abstract Schematic presentation of a colorimetric aptamer biosensor for adenosine detection based on DNA cycling amplification and salt-induced aggregation of gold nanoparticles.

     

Electrochemiluminescent aptasensor for thrombin using nitrogen-doped graphene quantum dots

An electrochemiluminescent (ECL) aptamer based method is described for the determination of thrombin. Three-dimensional nitrogen-doped graphene oxide (3D-NGO) was placed on a glassy carbon electrode (GCE) to provide an electrode surface that displays excellent electrical conductivity and acts as a strong emitter of ECL. The modified electrode was further coated with chitosan via electrodeposition. Finally, the amino-modified aptamer was immobilized on the modified GCE. The interaction between thrombin and aptamer results in a decrease in ECL. The assay has a linear response in the 1 fM to 1 nM thrombin concentration range and a 0.25 fM lower detection limit (at an S/N ratio of 3). The method was applied to the determination of thrombin in spiked human plasma samples, and recoveries ranged between 94 and 105% (with RSDs of <3.6%). The calibration plot was recorded at potential and wavelength of fluorescence emission (wavelength:?445 nm; potential:?0 to -2 V).

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Graphical abstract A bare glassy carbon electrode (GCE) does not display electrochemiluminescence (ECL). If, however, nitrogen-doped graphene quantum dots, chitosan, and three-dimensional nitrogen-doped graphene oxide (NGQD-chitosan/3D-NGO) are electrodeposited on the GCE, strong ECL can be observed. The ECL intensity decreased after aptamer and bovine serum albumin (BSA) were dropped onto the electrode (curve a). However, the ECL further decreases after addition of thrombin (TB; curve b).

     

Fluorescent quantum dots for microbial imaging

QDs (Semiconductor QDs, CDs, SiQDs, and Pdots) are used in imaging microorganisms including viruses, bacteria, and fungi.     

Aptasensor for visual and fluorometric determination of lysozyme based on the inner filter effect of gold nanoparticles on CdTe quantum dots

The authors describe an aptasensor for visual and fluorescent detection of lysozyme via an inner filter effect (IFE). The assay is based on the fact that red gold nanoparticles (AuNPs) act as powerful absorbers of the green fluorescence of CdTe because of spectral overlap. If the lysozyme-binding aptamer is adsorbed onto the surface of the AuNPs, the salt-induced aggregation of AuNPs (that leads to a color change from red to blue) does not occur and the IFE remains efficient. If lysozyme is present, it will bind the aptamer and thereby prevent its adsorption on the AuNPs. As a result, the salt-triggered aggregation of the AuNPs will occur. Consequently, color will change from red to blue, and green fluorescence will pop up because the IFE is suppressed. Under optimum conditions, fluorescence is linearly related to lysozyme concentration in the 1.0 nM to 20 nM concentration range, with a 0.55 nM limit of detection. The method is perceived to be of wider applicability in that it may be used to design other visual and fluorescent assays if appropriate aptamers are available.

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Graphical abstract The fluorescence intensity of QDs is quenched by gold nanoparticles (AuNPs) due to an inner filter effect. Aptamers can adsorb on AuNPs to prevent the salt-induced aggregation. AuNPs serve a dual function as fluorescence quencher and colorimetric reporter.

     

Mass-amplifying quantum dots in a fluorescence polarization-based aptasensor for ATP

We report on a fluorescence polarization assay for the detection of the target analyte ATP by making use of an aptasensor and of mass-amplifying CdTe-CdS quantum dots. The ATP aptamer was modified with digoxin antigen and hybridized with its complementary DNA that was modified with the CdTe-CdS quantum dots. Following the addition of digoxin antibody, the mass-amplifying aptasensor probe is formed as a result of the immuno reaction. In the presence of ATP, the polarization of fluorescence decreases because the digoxin antibody becomes dissociated due to the recognition of the ATP by the ATP aptamer. Under optimized conditions, the method has a linear response to ATP in the 10 to 350 μM concentration range, and the limit of detection is 3.7 μM. The method combines the specific recognition capability of aptamers with the sensitivity of an immunoreaction. It has good selectivity and sensitivity, and can be used to detect ATP in serum samples.

Electrochemical synthesis of polyaniline nanocomposite based on modified gold nanoparticles and its application for electrochemical aptasensor

Journal of Solid State Electrochemistry - The method of synthesis of polyaniline, as an important conductive polymer, and the conjugated metal nanomaterials applied in the polymer, determines the...     

An ultrasensitive quantum dots fluorescent polarization immunoassay based on the antibody modified Au nanoparticles amplifying for the detection of adenosine triphosphate

In this work, an ultrasensitive fluorescent polarization immunoassay (FPIA) method based on the quantum dot/aptamer/antibody/gold nanoparticles ensemble has been developed for the detection of adenosine triphosphate (ATP). DNA hybridization is formed when ATP is present in the PBS solution containing the DNA-conjugated quantum dots (QDs) and antibody-AuNPs. The substantial sensitivity improvement of the antibody-AuNPs-enhanced method is mainly attributed to the slower rotation of fluorescent unit when QDs-labeled oligonucleotides hybridize with antibody modified the gold nanoparticle. As a result, the fluorescent polarization (FP) values of the system increase significantly. Under the optimal conditions, a linear response with ATP concentration is ranged from 8 × 10⁻¹² M to 2.40 × 10⁻⁴ M. The detection limit reached as low as 1.8 pM. The developed work provides a sensitive and selective immunoassay protocol for ATP detection, which could be applied in more bioanalytical systems.     

Visual and fluorescent detection of acetamiprid based on the inner filter effect of gold nanoparticles on ratiometric fluorescence quantum dots

In this work, we develop a simple and rapid sensing method for the visual and fluorescent detection of acetamiprid (AC) based on the inner-filter effect (IFE) of gold nanoparticles (AuNPs) on ratiometric fluorescent quantum dots (RF-QDs). The RF-QDs based dual-emission nanosensor was fabricated by assembling green emissive QDs (QDs_539 nm, λ_em = 539 nm) on the surface of red emissive QDs (QDs_661 nm, λ_em = 661 nm)-doped silica microspheres. The photoluminescence (PL) intensity of RF-QDs could be quenched by AuNPs based on IFE. Acetamiprid can adsorb on the surface of AuNPs due to its cyano group that has good affinity with gold, which could induce the aggregation of AuNPs accompanying color change from red to blue. Thus, the IFE of AuNPs on RF-QDs was weakened and the PL intensity of RF-QDs was recovered accordingly. Under the optimized conditions, the PL intensity of the RF-QDs/AuNPs system was proportional to the concentration of AC in the range of 0.025–5.0 μg mL⁻¹, with a detection limit of 16.8 μg L⁻¹. The established method had been used for AC detection in environmental and agricultural samples with satisfactory results.     

Impedimetric aptasensor for Staphylococcus aureus based on nanocomposite prepared from reduced graphene oxide and gold nanoparticles

We describe here an aptasensor for the ultrasensitive detection of Staphylococcus aureus by electrochemical impedance spectroscopy (EIS). Single-stranded DNA was linked to a nanocomposite prepared from reduced graphene oxide (rGO) and gold nanoparticles (AuNP). Thiolated ssDNA was covalently linked to the AuNPs linked to rGO, and probe DNA was immobilized on the surface of an AuNP-modified glassy carbon electrode to capture and concentrate Staph. aureus. The probe DNA of the aptasensor selectively captures the target bacteria in its three-dimensional space, and these results in a dramatic increase in impedance. Scanning electron microscopy, cyclic voltammetry and EIS were used to monitor the single steps of the electrode assembly process. The effect was utilized to quantify the bacteria in the concentration range from 10 to 10⁶ cfu mL^?1 and with a detection limit of 10 cfu mL^?1 (S/N?=?3). The relative standard deviation of Staphylococcus aureus detection was equal to 4.3 % (10⁵ cfu mL^?1, n?=?7). In addition to its sensitivity, the biosensor exhibits high selectivity over other pathogens.

Binary heterogeneous superlattices assembled from quantum dots and gold nanoparticles with DNA

Controllable assembly of three-dimensional (3D) superlattices composed of different types of nanoscale objects opens new opportunities for material fabrication. Herein we show the successful assembly of heterogeneous 3D structures from gold nanoparticles (AuNPs) and quantum dots (QDs) using DNA encoding. By applying synchrotron-based small-angle X-ray scattering, we found that AuNPs and QDs are positioned in a body-centered cubic lattice, while each particle type, AuNP and QD, is arranged in a simple-cubic manner. Our studies demonstrate a route for assembly of integrated heterogeneous 3D structures from different nano-objects by DNA-encoded interactions.     

Multifunctional compact zwitterionic ligands for preparing robust biocompatible semiconductor quantum dots and gold nanoparticles

We describe the synthesis of a series of four different ligands which are used to prepare hydrophilic, biocompatible luminescent quantum dots (QDs) and gold nanoparticles (AuNPs). Overall, the ligands are designed to be compact while still imparting a zwitterionic character to the NPs. Ligands are synthesized appended to a bidentate dihydrolipoic acid- (DHLA) anchor group, allowing for high-affinity NP attachment, and simultaneously incorporate tertiary amines along with carboxyl and/or hydroxyl groups. These are placed in close proximity within the ligand structure and their capacity for joint ionization imparts the requisite zwitterionic nature to the nanocrystal. QDs functionalized with the four different compact ligands were subjected to extensive physical characterization including surface charge, wettability, hydrodynamic size, and tolerance to a wide pH range or high salt concentration over time. The utility of the compact ligand coated QDs was further examined by testing of direct conjugation to polyhistidine-appended protein and peptides, aqueous covalent-coupling chemistry, and the ability to engage in F?rster resonance energy transfer (FRET). Conjugating cell penetrating peptides to the compact ligand coated QD series facilitated their rapid and efficient cellular uptake, while subsequent cytotoxicity tests showed no apparent decreases in cell viability. In vivo biocompatibility was also demonstrated by microinjecting the compact ligand coated QDs into cells and monitoring their stability over time. Inherent benefits of the ligand design could be extended beyond QDs as AuNPs functionalized with the same compact ligand series showed similar colloidal properties. The strong potential of these ligands to expand NP capabilities in many biological applications is highlighted.     

Electrochemical aptasensor for detection of copper based on a reagentless signal-on architecture and amplification by gold nanoparticles

A highly sensitive and specific electrochemical aptasensor for Cu(2+) detection based on gold nanoparticles (AuNPs) is presented. In this work, AuNPs offered a big surface area to immobilize a large number of aptamers and excellent electrochemical signal transduction. Its high sensitivity, low detection limit, and wide detection range are the main advantages over our former copper aptasensor. The peak current increased proportionally to the Cu(2+) concentration over the range from 0.1 nM to 10 μM with a detection limit of 0.1 pM. The presence of other divalent metal ions did not affect the detection of Cu(2+), which indicates a high specificity of Cu(2+) detection could be detected. Rapidity, simplicity, and excellent selectivity make it suitable for practical use in determination of Cu(2+) from lake samples.     

A high-throughput homogeneous immunoassay based on Förster resonance energy transfer between quantum dots and gold nanoparticles

A novel homogeneous immunoassay based on Förster resonance energy transfer for sensitive detection of tumor, e.g., marker with carcinoembryonic antigen (CEA), was proposed. The assay was consisted of polyclonal goat anti-CEA antibody labeled luminescent CdTe quantum dots (QDs) as donor and monoclonal goat anti-CEA antibody labeled gold nanoparticles (AuNPs) as acceptor. In presence of CEA, the bio-affinity between antigen and antibody made the QDs and AuNPs close enough, thus the photoluminescence (PL) quenching of CdTe QDs occurred. The PL properties could be transformed into the fluorometric variation, corresponding to the target antigen concentration, and could be easily monitored and analyzed with the home-made image analysis software. The fluorometric results indicated a linear detection range of 1–110 ng mL⁻¹ for CEA, with a detection limit of 0.3 ng mL⁻¹. The proposed assay configuration was attractive for carcinoma screening or single sample in point-of-care testing, and even field use. In spite of the limit of available model analyte, this approach could be easily extended to detection of a wide range of biomarkers.     

The quenching of CdSe quantum dots photoluminescence by gold nanoparticles in solution

The photoluminescence (PL) of CdSe quantum dots (QD) in aqueous media has been studied in the presence of gold nanoparticles (NP) with different shapes. The steady state PL intensity of CdSe QD (1.5-2 nm in size) is quenched in the presence of gold NP. Picosecond bleach recovery and nanosecond time-resolved luminescence measurements show a faster bleach recovery and decrease in the lifetime of the emitting states of CdSe QD in the presence of quenchers. Surfactant-capped gold nanorods (NR) with aspect ratio of 3 and surfactant-capped and citrate-capped nanospheres (NS) of 12 nm diameter were used as quenchers in order to study the effect of shape and surface charge on the quenching rates. The Stern-Volmer kinetics model is used to examine the observed quenching behavior as a function of the quencher concentration. It was found that the quenching rate of NR is more than 1000 times stronger than that of NS with the same capping material. We also found that the quenching rate decreases as the length of the NR decreases, although the overlap between the CdSe emission and the NR absorption increases. This suggests that the quenching is a result of electron transfer rather than long-range (Forster-type) energy transfer processes. The quenching was attributed to the transfer of electron with energies below the Fermi level of gold to the trap holes of CdSe QD. The observed large difference between NR and NS quenching efficiencies was attributed to the presence of the [110] facets only in the NR, which have higher surface energy.     

Inhibition assay of biomolecules based on fluorescence resonance energy transfer (FRET) between quantum dots and gold nanoparticles

An inhibition assay method was developed based on the modulation in the FRET efficiency between quantum dots (QDs) and gold nanoparticles (AuNPs) in the presence of the molecules which inhibit the interactions between QD- and AuNP-conjugated biomolecules. For the functionalization, AuNPs were first stabilized by chemisorption of n-alkanethiols and then capped with the first generation polyamidoamine (G1 PAMAM) dendrimers. By employing a streptavidin-biotin couple as a model system, avidin was quantitatively analyzed as an inhibitor by sensing the change in photoluminescence (PL) quenching of SA-QDs by biotin-AuNPs. The detection limit for avidin was about 10 nM. It is anticipated that the PL quenching-based sensing system can be used for the quantitative analysis and high throughput screening of molecules which inhibit the specific biomolecular interactions.     

Amplified impedimetric aptasensor based on gold nanoparticles covalently bound graphene sheet for the picomolar detection of ochratoxin A

An amplified electrochemical impedimetric aptasensor for ochratoxin A (OTA) was developed with picomolar sensitivity. A facile route to fabricate gold nanoparticles covalently bound reduced graphene oxide (AuNPs–rGO) resulted in a large number of well-dispersed AuNPs on graphene sheets with tremendous binding sites for DNA, since the single rGO sheet and each AuNP can be loaded with hundreds of DNA strands. An aptasensor with sandwich model was fabricated which involved thiolated capture DNA immobilized on a gold electrode to capture the aptamer, then the sensing interface was incubated with OTA at a desired concentration, followed by AuNPs–rGO functionalized reporter DNA hybridized with the residual aptamers. By exploiting the AuNPs–rGO as an excellent signal amplified platform, a single hybridization event between aptamer and reporter DNA was translated into more than 10⁷ redox events, leading to a substantial increase in charge-transfer resistance (R_ct) by 7∼ orders of magnitude compared with that of the free aptamer modified electrode. Such designed aptasensor showed a decreased response of R_ct to the increase of OTA concentrations over a wide range of 1 pg mL⁻¹–50 ng mL⁻¹ and could detect extremely low OTA concentration, namely, 0.3 pg mL⁻¹ or 0.74 pM, which was much lower than that of most other existed impedimetric aptasensors. The signal amplification platform presented here would provide a promising model for the aptamer-based detection with a direct impedimetric method.     

Influence of gold nanoparticles on nonradiative energy transfer in nanoclusters of colloidal quantum dots InP@ZnS

The influence of gold nanoparticles on the efficiency of nonradiative resonance energy transfer between InP@ZnS quantum dots composed of nanoclusters, a new type of organized structures consisting of hydrophobic quantum dots and gold nanoparticles, has been experimentally studied.