And Unmodified Gold Nanoparticles Research Articles

A simple and sensitive AuNPs-based colorimetric aptasensor for specific detection of azlocillin

A new colorimetric biosensor for specific detection of azlocillin was developed by using DNA aptamer as recognition element and unmodified gold nanoparticles (AuNPs) as colorimetric indicator. In the absence of azlocillin, the AuNPs were protected by the aptamer and stabilized at high NaCl concentrations, displaying a red solution. In the presence of azlocillin, the aptamer reacts specifically with azlocillin, resulting in the aggregation of AuNPs and an apparent red to blue color change. The characteristic change can be easily observed by the naked eye and quantitatively detected by an ultraviolet–visible (UV–Vis) spectrometer. Under the optimal conditions, the absorbance variation at 522 nm (ΔA522) of AuNPs changed proportionally with increasing concentration of azlocillin, which exhibited a linear relationship in the concentration range of 50 nM to 500 nM, with a detection limit of 11.6 nM. Furthermore, the aptasensor was successfully used to detect azlocillin in milk and tap water samples, with recoveries ranging from 97.64% to 102.21% and a relative standard deviation (RSD) less than 3.81%.

Label-free peptide nucleic acid biosensor for visual detection of multiple strains of influenza A virus suitable for field applications

Accurate and rapid diagnosis of Influenza A viruses (IAVs) is challenging because of multiple strains circulating in humans and animal populations, and the emergence of new strains. In this study, we demonstrate a simple and rapid strategy for visual detection of multiple strains of IAVs (H1 to H16 subtypes) using peptide nucleic acid (PNA) as a biosensor and unmodified gold nanoparticles (AuNPs) as a reporter. The design principle of the assay is based on the color change on account of free PNA-induced aggregation of AuNPs in the presence of non-complementary viral RNA sequence and vice-versa. The assay could detect IAV RNA with a visual limit of detection of 2.3 ng. The quantification of RNA with a considerable accuracy on a simple spectrophotometer was achieved on plotting the PNA-induced colorimetric changes (absorption ratio of A640/A520) in the presence of a varying concentration of complementary RNA. As a proof-of-concept, the visual assay was validated on 419 avian clinical samples and receiver operating characteristic (ROC) curve analysis showed a high diagnostic specificity (96.46%, 95% CI = 93.8 to 98.2) and sensitivity (82.41%, 95% CI = 73.9 to 89.1) when RT-qPCR was used as reference test. Hence, the simplicity, rapidity, and universality of this strategy make it a potential candidate visual assay for clinical diagnosis and surveillance of IAVs, especially in the resource-limited settings. The proposed strategy establishes new avenues for developing a simple and rapid diagnostic system for viral infections and biomolecules.

Highly Sensitive Colorimetric Biosensor for Staphylococcal Enterotoxin B by a Label-Free Aptamer and Gold Nanoparticles.

A simple, sensitive and selective colorimetric biosensor for the detection of Staphylococcal enterotoxin B (SEB) was developed using SEB-binding aptamer (SEB2) as recognition element and unmodified gold nanoparticles (AuNPs) as colorimetric probes. The assay is based on color change from red to purple due to conformational change of aptamer in the presence of SEB, and the phenomenon of salt-induced AuNPs aggregation which could be monitored by naked eye or UV–vis spectrometer. Results showed that the AuNPs can effectively differentiate the SEB induced conformational change of the aptamer in the presence of a given high salt concentration. A linear response in the range of 50 μg/mL to 0.5 ng/mL of SEB concentration was obtained. The assay was highly specific to SEB as compared to other related toxins. The limit of detection (LOD) of SEB achieved within few minutes was 50 ng/mL visually and spectrometric method improved it to 0.5 ng/mL. Robustness of the assay was tested in artificially spiked milk samples and cross-checked using in house developed sandwich ELISA (IgY as capturing and SEB specific monoclonal as revealing antibody) and PCR. This colorimetric assay could be a suitable alternative over existing methods during biological emergencies due to its simplicity, sensitive and cost effectiveness.

Asymmetric polymerase chain assay combined with propidium monoazide treatment and unmodified gold nanoparticles for colorimetric detection of viable emetic Bacillus cereus in milk

Bacillus cereus is a causative agent of an emetic food-borne disease. In this study, visual detection of viable emetic B. cereus was developed using a propidium monoazide (PMA)-asymmetric polymerase chain reaction (asPCR) and unmodified gold nanoparticles (AuNPs). To detect only the viable emetic B. cereus, PMA treatment was selected before DNA extraction to eliminate the false-positive results from dead bacteria. In the presence of viable target bacteria, the long genomic DNA fragments from the cereulide synthetase gene (cesB) were produced by asPCR, which could be effectively absorbed onto naked AuNPs via coordination between Au and the nitrogen atoms of the exposed bases. After adding NaCl solution, visual detection with the naked-eye or with UV–vis spectrophotometer was possible within a few minutes. Under the optimum conditions, the limit of detection (LOD) for viable emetic B. cereus reached as low as 9.2×101 CFU/mL in 0.01M phosphate-buffered saline and 3.4×102 CFU/mL in milk, which was adequate to meet the maximum limit imposed by the Commission Regulation (EC) No 2073/2005 (500 CFU/mL). The proposed method also exhibited excellent discrimination against 10 common pathogenic bacteria in milk. The PMA-asPCR-AuNPs colorimetric assay offers a promising application in the detection of low concentrations of food-borne pathogens.

A simple and sensitive aptasensor for colorimetric detection of adenosine triphosphate based on unmodified gold nanoparticles

A simple and sensitive colorimetric aptasensor for rapid and facile detection of adenosine triphosphate (ATP) has been demonstrated here based on aptamer-based hairpin probes and unmodified gold nanoparticles (AuNPs). The hairpin probe is constructed by adding another five nucleotides to the 5′-end of an anti-ATP aptamer which can hybridize to nucleotides at the 3′-end of the aptamer, forming a hairpin-shaped structure. In the absence of ATP, the hairpin probes are rigid, and the AuNPs are susceptible to salt-induced aggregation. Conversely, upon binding with target ATP, the hairpin probes undergo conformational changes, forming aptamer-ATP complexes and exposing flexible ends which coat the surface of AuNPs to inhibit their aggregation in the high salt solution. Subsequently, a blue-to-red color change can be recognized by the naked eye. The aptasensor achieved selective responses toward ATP with a detection limit of 0.1μM, and exhibited high-quality detection performance in biological samples. In addition, this detection method is simple, rapid and cost-effective, holding great potential for further applications in point-of-care research.

Utilization of unmodified gold nanoparticles for label-free detection of mercury (II): Insight into rational design of mercury-specific oligonucleotides

Colorimetric detection of mercury (II) with the use of DNA oligonucleotides and unmodified gold nanoparticles (AuNPs) as indicators has been extensively studied. This study provides in-depth insights into the rational design of mercury-specific oligonucleotides (MSO) in the biosensing system. The leftover bases of MSO, as a result of the formation of T–Hg2+–T base pairs, can adsorb on the AuNPs and hinder their aggregation at concentrations of salt. This phenomenon was directly verified by the changes in particle sizes characterized by dynamic light scattering for the first time. Based on these findings, we proposed a rational design for the MSO with approximately 20-fold improvement in detection sensitivity. The detection limit of the proposed assay decreased to 15nM with a linear working range from 50nM to 300nM for Hg2+. The cross-reactivity against eight other metal ions was negligible compared with the response to Hg2+. Considering the diverse applications of AuNPs with oligonucleotides, this study can serve as a good reference and provides important implications in sensing and DNA-directed nanoparticle assembly.

A label-free fluorescence assay for thrombin activity analysis based on fluorescent protein and gold nanoparticles

A label-free and sensitive fluorescence assay has been developed for probing thrombin activity based on an engineered enhanced green fluorescent protein (EGFP) probe and unmodified gold nanoparticles (AuNPs).

Label-free colorimetric detection of Cu2+ on the basis of Fenton reaction-assisted signal amplification with unmodified gold nanoparticles as indicator

A novel label-free strategy is demonstrated for the amplified detection of copper ions (Cu2+) on the basis of Fenton reaction and unmodified gold nanoparticles (AuNPs). AuNPs are known to stabilise in the presence of single-stranded DNA (ssDNA), which prevents the salt-induced aggregation of AuNPs in solution because of electrostatic repulsion. However, in the presence of a Fenton reagent, Cu2+ catalyses the Fenton reaction to generate hydroxyl free radicals, which cut the ssDNA into different sequence fragments or even single bases to induce AuNP aggregation along with a red-to-blue colour change. Cu2+ is qualitatively and quantitatively detected through this phenomenon, and Cu2+ recycling may amplify the signal transduction based on Fenton reaction with unlabelled sensing strategies. A linear calibration of A700/A525 versus Cu2+ concentration is obtained within 0.1–10μM, along with a detection limit of 23nM and relative standard deviation (RSD) of 4.1% (1μM). Moreover, the sensing system exhibits excellent sensitivity and selectivity for Cu2+, with quantitative spike-recoveries from 88% to 116% in environment water samples.

Determination of Tigecycline by Quantum Dots/Gold Nanoparticles-based Fluorescent and Colorimetric Sensing System

In this study, two simple and sensitive methods for detection of tigecycline are developed based on thioglycolic acid-capped CdTe quantum dots (TGA-CdTe QDs) and unmodified gold nanoparticles (AuNPs), respectively. The first method was based on the fluorescence quenching of CdTe QDs by the addition of tigecycline. For the second method, the introduction of tigecycline could induce the aggregation of gold nanoparticles, displaying distinct changes in color and in UV-vis spectra. Under optimal conditions, the relative fluorescence intensity of CdTe QDs decreased linearly with the increasing concentration of tigecycline in the range of 0.11-35.56 ug mL-1 with a detection limit of 3.93×10-2 ug mL-1. While the colorimetric sensor could detect tigecycline in linear range from 5.98×10-3 to 1.44×10-1 ug mL-1 and from 0.30 to 2.99 ug mL-1 with a detection limit of 1.07×10-3 ug mL-1. Moreover, both the proposed methods were applied to real sample with satisfactory results. Finally, comparisons between these two methods were made. Keywords: Colorimetric, fluorescence quenching, gold nanoparticles, quantum dots, tigecycline.

AuNPs-Based Colorimetric Assay for Identification of Chicken Tissues in Meat and Meat Products

A simple colorimetric assay was developed to identify chicken tissues in meat and meat products by utilizing thiol-labeled primers and unmodified gold nanoparticles (AuNPs). Primers were designed based on the chicken-specific mitochondrial D-loop gene. Polymerase chain reaction (PCR) is applied to amplify the target gene, and the PCR products labeled with thiol at one end were obtained. Following the mixing of AuNPs with the PCR products, the thiol binds to the surface of AuNPs, resulting in the formation of GNP-PCR products. The resultant PCR products had abundant negative charges, which made AuNPs maintain dispersion under the role of electrostatic repulsion. As a result, in the presence of PCR products, AuNPs remained red in the presence of salt. In the absence of PCR products, the color of AuNPs changed from red to blue; therefore, the method described here could be exploited for the verification of chicken tissues with high accuracy.

A dual-mode colorimetric and fluorometric "light on" sensor for thiocyanate based on fluorescent carbon dots and unmodified gold nanoparticles.

A novel, highly sensitive and selective dual-readout (colorimetric and fluorometric) sensor based on fluorescent carbon dots (CDs) and unmodified gold nanoparticles (AuNPs) for the detection of thiocyanate (SCN(-)) was proposed. Amino-functionalized CDs could be readily adsorbed onto the surface of citrate-stabilized AuNPs through Au-N interactions, leading to the aggregation of AuNPs and the nonfluorescent off-state of CDs arising from potential fluorescence resonance energy transfer (FRET). However, SCN(-) had a stronger affinity toward AuNPs and could compete with CDs to bind onto the surface of AuNPs in priority, which prevented the aggregation of AuNPs and fluorescence quenching of CDs. Correspondingly, both the colorimetric and fluorometric signals remained "light-on". The color of the sensing solution remained red and the fluorescence remained unquenched. A distinguishable change in the color was observed at a SCN(-) concentration of 1 μM by the naked eye and a detection limit as low as 0.036 μM was obtained by virtue of fluorescence spectroscopy. Both colorimetric and fluorometric sensors exhibited excellent selectivity toward SCN(-) over other common metallic ions and anions. In addition, such a sensing assay featured simplicity, rapidity, cost-effectiveness and ease of operation without further modification. The accuracy and precision were evaluated based on the quantitative detection of SCN(-) in tap water and saliva samples with satisfactory results.

A simple “clickable” biosensor for colorimetric detection of copper(II) ions based on unmodified gold nanoparticles

A novel colorimetric copper(II) biosensor has been developed based on the high specificity of alkyne–azide click reaction to the catalysis of copper ions and unmodified gold nanoparticles (AuNPs) as the signal reporter. The clickable DNA probe consists of two parts: an azide group-modified double-stranded DNA (dsDNA) hybrid with an elongated tail and a short alkyne-modified single-stranded DNA (ssDNA). Because of low melting temperature of the short ssDNA, these two parts are separated in the absence of Cu2+. Copper ion-induced azide–alkyne click ligation caused a structural change of probe from the separated form to entire dsDNA form. This structural change of probe can be monitored by the unmodified AuNPs via mediating their aggregation with a red-to-blue colorimetric read-out because of the differential ability of ssDNA and dsDNA to protect AuNPs against salt-induced aggregation. Under the optimum conditions, this biosensor can sensitively and specifically detect Cu2+ with a low detection limit of 250nM and a linear range of 0.5–10μM. The method is simple and economic without dual-labeling DNA and AuNPs modification. It is also highly selective for Cu2+ in the presence of high concentrations of other environmentally relevant metal ions because of the great specificity of the copper-caused alkyne–azide click reaction, which potentially meets the requirement of the detection in real samples.

Colorimetric detection of apoptosis based on caspase-3 activity assay using unmodified gold nanoparticles

A new label-free method for the detection of apoptosis was proposed based on colorimetric assay of caspase-3 activity using an unlabeled Asp-Glu-Val-Asp (DEVD)-containing peptide substrate and unmodified gold nanoparticles (AuNPs).

Colorimetric multiplexed analysis of mercury and silver ions by using a unimolecular DNA probe and unmodified gold nanoparticles

In this paper, a colorimetric assay for multiplexed analysis of mercury and silver ions was demonstrated by using a rationally designed unimolecular multifunctional DNA probe (UMDP) as sensing element, and unmodified gold nanoparticles (AuNPs) as color-reporting probes. The UMDP had a random coil structure that changed into two different hairpin-like structures with a T–Hg2+–T or C–Ag+–C basepairing built-in the stem upon binding Hg2+ or Ag+ ions, respectively. As a result, the conformation changes facilitated the salt-induced AuNPs aggregation, leading red-to-blue color change with a red shift of the plasmon band in the UV-visible absorption spectrum. Thus, the change in absorption intensity allowed the multiplexed detection of Hg2+ and Ag+ ions. The sensing system could detect as low as 2.5 × 10−7 M Hg2+ ions and 5 × 10−7 M Ag+ ions. Both the color and absorption changes of the system were selective for Hg2+ and Ag+ ions, which met the selective requirements for biomedical and environmental application. Moreover, the two target ions were simply distinguished by using EDTA, Hg2+ ions could not cause AuNPs aggregation while Ag+ ions still could in the presence of EDTA. Then, the colorimetric response of the two target ions in spiked lake water and tap water were tested, respectively. We expect this simple and cost-effective method to have wide-ranging applications for monitoring water quality in the developing region.

Aptamer-based colorimetric biosensing of dopamine using unmodified gold nanoparticles

A simple, sensitive and selective colorimetric biosensor for the detection of dopamine (DA) was demonstrated with a 58-mer dopamine-binding aptamer (DBA) as recognition element and unmodified gold nanoparticles (AuNPs) as probes. Upon the addition of DA, the conformation of DBA would change from a random coil structure to a rigid tertiary structure like a pocket and this change has been demonstrated by circular dichroism spectroscopic experiments. Besides, the conformational change of DBA could facilitate salt-induced AuNP aggregation and lead to the color change of AuNPs from red to blue. The calibration modeling showed that the analytical linear range covered from 5.4 × 10 −7 M to 5.4 × 10 −6 M and the corresponding limit of detection (LOD) was 3.6 × 10 −7 M. Some common interferents such as 3,4-dihydroxyphenylalanine (DOPA), catechol, epinephrine (EP), 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), and ascorbic acid (AA) showed no or just a little interference in the determination of DA.