A rapid and simple ratiometric fluorescent sensor for patulin detection based on a stabilized DNA duplex probe containing less amount of aptamer-involved base pairs.

In this study, a sensor is described for determination of patulin by using ratiometric fluorescence measurement and strand displacement strategy. In the presence of patulin, the ratiometric fluorescence response decreases, owing to disassembly of DNA duplex structure and target-mediated release of TAMRA-labeled complementary DNA sequence2 (cDNA2). While, in the absence of target, the fluorescence resonance energy transfer (FRET) phenomenon happens between FAM and TAMRA under excitation at 490 nm, resulting in the enhancement of ratiometric signal. The use of ratiometric fluorescence signal with different signal indicators avoids the problem of environmental interference and improves the sensitivity of the aptasensor. Also, the DNA duplex structure contains minimum aptamer-involved base pair sequence, resulting in further improvement of the aptasensor sensitivity. This sensing platform provided a wide linear range from 15 ng/L to 35 µg/L and a detection limit of 6 ng/L for patulin. The aptasensor was used to determine patulin in spiked apple juice samples and showed satisfactory results.

A novel electrochemical aptasensor based on nontarget-induced high accumulation of methylene blue on the surface of electrode for sensing of α-synuclein oligomer.

This study describes a novel electrochemical aptasensor for detection of α-synuclein (α-syn) oligomer, an important biomarker related to Parkinson's and Alzheimer's diseases. The sensing platform is based on exonuclease I (Exo I), terminal deoxynucleotidyl transferase (TdT) and methylene blue. The aptasensor exploits the improved sensitivity because of applications of TdT and Exo I and also a label-free aptamer (Apt). Furthermore, direct immobilization of complementary strand of aptamer (CS) instead of Apt on the surface of electrode prohibits Apt self-assembled monolayer aggregation and keeps the function of the Apt. In the absence of α-syn oligomer, TdT enhances lengths of Apt and CS and so, increases accumulation of methylene blue as redox agent on the surface of electrode, leading to a strong current signal. While in the presence of α-syn oligomer, Exo I digests CS on the electrode surface, resulting in less accumulation of methylene blue on the electrode surface and a weak current signal. The relative electrochemical signal of the aptasensor increased linearly with the logarithm of α-syn oligomer concentration in the range from 60 pM to 150 nM. The detection limit was 10 pM. Furthermore, the sensor showed high precision and repeatability for detection of α-syn oligomer in serum samples.

A novel colorimetric aptasensor for ultrasensitive detection of cocaine based on the formation of three-way junction pockets on the surfaces of gold nanoparticles.

Herein, a novel colorimetric aptasensor was introduced for detection of cocaine based on the formation of three-way junction pockets on the surfaces of gold nanoparticles (AuNPs) and the catalytic activity of the surfaces of AuNPs. Simplicity and detection of cocaine in a short time (only 35 min) are some of the unique features of the proposed sensing strategy. In the presence of cocaine, triple-fragment aptamer (TFA) forms on the surfaces of AuNPs, leading to a significant decrease of the catalytic activity of AuNPs and the color of samples remains yellow. In the absence of target, TFA does not form on the surfaces of AuNPs and 4-Nitrophenol, as a colorimetric agent, has more access to the surfaces of AuNPs, resulting in the reduction of 4-Nitrophenol and the color of sample changes from yellow to colorless. The sensing strategy showed good specificity, a limit of detection (LOD) of 440 pM and a dynamic range over 2-100 nM. The sensing method was also successfully applied to detect cocaine in spiked human serum samples with recovery of 94.71-98.63%.

Novel Colorimetric Aptasensor for Zearalenone Detection Based on Nontarget-Induced Aptamer Walker, Gold Nanoparticles, and Exonuclease-Assisted Recycling Amplification.

Zearalenone (ZEN) toxicity is a significant risk for human beings. Thus, it is of high importance to develop sensitive, precise, and inexpensive analytical methods for ZEN detection, especially in human serum. Here, a colorimetric aptasensor is presented for the determination of ZEN based on the nontarget-induced aptamer walker, catalytic reaction of gold nanoparticles (AuNPs), exonuclease III (Exo III) as a signal amplifier, and 4-nitrophenol as a colorimetric agent. Low amount of ZEN requirement and signal amplification are some of the distinct advantages of the proposed aptasensor. In the absence of ZEN, the aptamer (Apt) starts walking on the AuNP surface with the help of Exo III and binds to multiple complementary strands of aptamer, leading to the change of sample color from yellow to colorless. Upon the addition of ZEN, both the Apt and complementary strand exist as single-stranded DNAs on the surface of AuNPs, resulting in less access of 4-nitrophenol to the surface of AuNPs and less catalytic performance of AuNPs. In this situation, the color of the sample remains yellow (the color of 4-nitrophenol). The presented aptasensor was capable to detect ZEN in a wide linear dynamic range, 20-80 000 ng/L, with a detection limit of 10 ng/L. The prepared sensing strategy was successfully used for ZEN determination in the human serum sample.

A colorimetric gold nanoparticle aggregation assay for malathion based on target-induced hairpin structure assembly of complementary strands of aptamer.

The authors describe a method for the colorimetric determination of the pesticide malathion. It is based on the use of a hairpin structure consisting of a complementary strand of aptamer and a double-stranded DNA (dsDNA) structure to protect gold nanoparticles (AuNPs) against salt-induced aggregation. In the absence of malathion, the dsDNA structure is preserved on the surface of AuNPs and the color of the AuNPs in solutions containing NaCl remains red. However, in the presence of malathion, a hairpin structure of complementary strand is formed. The Aptamer/Malathion complex and the complementary strand are released from the surface of the AuNPs. As a result, the AuNPs undergo salt-induced aggregation which is accompanied by a color change to blue. The assay allows malathion to be quantified within 35 min (A650/A520 was measured). The detection limit is 1 pM, and response is linear in the 5 pM to 10 nM malathion concentration range. The method is specific and was successfully applied to the determination of malathion in spiked human serum samples. Graphical abstract Schematic representation of detection of malathion based on dsDNA-modified gold nanoparticles (AuNPs) and the hairpin structure of the complementary strand.

Serum concentrations of MCP-1 and IL-6 in combination predict the presence of coronary artery disease and mortality in subjects undergoing coronary angiography.

Cytokines play a key role in the pathogenesis of coronary artery disease (CAD). The aim of current study was to investigate the relationship between the serum concentrations of 12 cytokines with mortality and extent of CAD in individuals undergoing angiography and healthy controls. 342 CAD patients were recruited and divided into 2 groups: those with ≥50% occlusion in at least one coronary artery [Angiography (+)] or <50% obstruction in coronary arteries [Angiography (-)]. Also 120 healthy subjects were enrolled as control group. Lipid profile, fasting blood glucose, body mass index, and blood pressure were evaluated in all the subjects. An Evidence Investigator® was used for measuring 12 cytokines (IL-1α, IL-1ß, IL-2, IL-4, IL-6, IL-8, IL-10, TNF-α, MCP-1, IFN-γ, EGF, VEGF) using sandwich chemiluminescent assays. Univariate analysis, multivariate regression models, ROC, and Kaplan-Meier survival curves were used for exploring the candidate markers in CAD patients. Serum level of IFN-γ, IL-4, MCP-1, EGF, IL-6, and IL-8 were markedly higher in angiogram-positive patients, while VEGF concentrations were significantly (P < 0.05) lower, compared to control group. ROC analysis for MCP-1 showed that a cut-off of 61.95 pg/mL had 91% sensitivity and 91% specificity for predicting CAD patients. Moreover, >2.16 pg/mL IL-6 had a > 94% sensitivity and 70% specificity in predicting 2 years mortality in the subjects with a serum MCP-1 > 61.95 pg/ mL, and patients having IL-6/MCP-1 combination had a shorter survival.Our findings demonstrate that CAD patients with serum MCP-1 and IL-6 levels of >61.95 and >2.16 pg/mL had a higher mortality with 94.1% sensitivity and 70.5% specificity for predicting mortality in CAD patients.

Double targeting, controlled release and reversible delivery of daunorubicin to cancer cells by polyvalent aptamers-modified gold nanoparticles.

Clinical use of daunorubicin (Dau) in treatment of leukemia has been restricted because of its cardiotoxicity. Targeted delivery of anticancer drugs could decrease their off-target effects and enhance their efficacy. In this study a modified polyvalent aptamers (PA)-Daunorubicin (Dau)-Gold nanoparticles (AuNPs) complex was designed and its efficacy was assessed in Molt-4 cells (human acute lymphoblastic leukemia T-cell, target). Dau was efficiently loaded (10.5 µM) onto 1mL of PA-modified AuNPs. Dau was released from the PA-Dau-AuNPs complex in a pH-sensitive manner (faster release at pH5.5). The results of flow cytometry analysis indicated that the PA-Dau-AuNPs complex was efficiently internalized into target cells, but not into nontarget cells. The results of MTT assay were consistent with the internalization data. PA-Dau-AuNPs complex had less cytotoxicity in U266 cells compared to Dau alone and even Apt-Dau-AuNPs complex. The PA-Dau-AuNPs complex had more cytotoxicity in Molt-4 cells compared to Dau alone and even Apt-Dau-AuNPs complex. Cytotoxicity of PA-Dau-AuNPs complex was effectively antagonized using antisense of polyvalent aptamers. In conclusion, the designed drug delivery system inherited the properties of efficient drug loading, tumor targeting, pH-dependent drug release and controllable delivery of Dau to tumor cells.

A novel electrochemical aptasensor based on Y-shape structure of dual-aptamer-complementary strand conjugate for ultrasensitive detection of myoglobin.

Monitoring of myoglobin (Mb) in human blood serum is highly in demand for early diagnosis of acute myocardial infarction (AMI). Here, a novel electrochemical aptasensor was developed for ultrasensitive and selective detection of Mb, based on Y-shape structure of dual-aptamer (DApt)-complementary strand of aptamer (CS) conjugate, gold electrode and exonuclease I (Exo I). The designed aptasensor obtains features of gold, such as high electrochemical conductivity and large surface area, property of Y-shape structure of DApt-CS conjugate to function as a gate and obstacle for the access of redox probe to the surface of electrode, as well as high specificity and sensitivity of aptamer toward its target and Exo I as an enzyme which specifically degrades the 3'-end of single-stranded DNA (ssDNA). In the absence of Mb, the Y-shape structure remains intact. So, a weak electrochemical signal is observed. Upon addition of target, the DApt leave the CS and bind to Mb, leading to disassembly of Y-shape structure and following the addition of Exo I, a strong electrochemical signal could be recorded. The fabricated aptasensor showed high selectivity toward Mb with a limit of detection (LOD) as low as 27 pM. Besides, the developed aptasensor was effectively applied to detect Mb in human serum.

A novel colorimetric sandwich aptasensor based on an indirect competitive enzyme-free method for ultrasensitive detection of chloramphenicol.

Analytical methods for detection and quantitation of chloramphenicol in blood serum and foodstuffs arse highly in demand. In this study, a colorimetric sandwich aptamer-based sensor (aptasensor) was fabricated for sensitive and selective detection of chloramphenicol, based on an indirect competitive enzyme-free assay using gold nanoparticles (AuNPs), biotin and streptavidin. The designed aptasensor acquires characteristics of AuNPs, including large surface area and unique optical properties, and strong interaction of biotin with streptavidin. In the absence of chloramphenicol, the sandwich structure of aptasensor forms, leading to the observation of sharp red color. In the presence of target, functionalized AuNPs could not bind to 96-well plates, resulting in a faint red color. The fabricated colorimetric aptasensor exhibited high selectivity toward chloramphenicol with a limit of detection as low as 451 pM. Moreover, the developed colorimetric aptasensor was successfully used to detect chloramphenicol in milk and serum with LODs of 697 and 601 pM, respectively.

A novel fluorescent aptasensor based on hairpin structure of complementary strand of aptamer and nanoparticles as a signal amplification approach for ultrasensitive detection of cocaine.

Cocaine is one of the most commonly misused stimulant which could influence the central nervous system. In this study, a fluorescent aptamer-based sensor (aptasensor) was designed for sensitive and selective detection of cocaine, based on hairpin structure of complementary strand of aptamer (CS), target-induced release of aptamer (Apt) from CS and two kinds of nanoparticles, including silica nanoparticles (SNPs) coated with streptavidin and gold nanoparticles (AuNPs). The designed aptasensor acquires characteristics of AuNPs such as unique optical properties and large surface area, SNPs as amplifiers of fluorescence intensity, higher affinity of Apt toward its target relative to its CS, and finally the hairpin structure of CS that brings the fluorophore (FAM) to close proximity to the surface of SNPs. In the absence of cocaine, FAM is in close proximity to the surface of AuNPs, resulting in a weak fluorescence emission. In the presence of target, FAM comes to close proximity to the surface of SNPs because of the formation of hairpin structure of CS, leading to a very strong fluorescence emission. The fabricated fluorescent aptasensor exhibited a good selectivity toward cocaine with a limit of detection (LOD) as low as 209 pM. Moreover, the designed aptasensor was successfully utilized to detect cocaine in serum with a LOD as low as 293 pM.

Colorimetric and fluorescence quenching aptasensors for detection of streptomycin in blood serum and milk based on double-stranded DNA and gold nanoparticles.

Antibiotic residues in animal foodstuffs are of great concern to consumers. In this study, fluorescence quenching and colorimetric aptasensors were designed for detection of streptomycin based on aqueous gold nanoparticles (AuNPs) and double-stranded DNA (dsDNA). In the absence of streptomycin, aptamer/FAM-labeled complementary strand dsDNA is stable, resulting in the aggregation of AuNPs by salt and an obvious color change from red to blue and strong emission of fluorescence. In the presence of streptomycin, aptamer binds to its target and FAM-labeled complementary strand adsorbs on the surface of AuNPs. So the well-dispersed AuNPs remain stable against salt-induced aggregation with a wine-red color and the fluorescence of FAM-labeled complimentary strand is efficiently quenched by AuNPs. The colorimetric and fluorescence quenching aptasensors showed excellent selectivity toward streptomycin with limit of detections as low as 73.1 and 47.6 nM, respectively. The presented aptasensors were successfully used to detect streptomycin in milk and serum.

A novel electrochemical aptasensor based on single-walled carbon nanotubes, gold electrode and complimentary strand of aptamer for ultrasensitive detection of cocaine.

Cocaine is a strong central nervous system stimulant and one of the most commonly abused drugs. In this study, an electrochemical aptasensor was designed for sensitive and selective detection of cocaine, based on single-walled carbon nanotubes (SWNTs), gold electrode and complimentary strand of aptamer (CS). This electrochemical aptasensor inherits properties of SWNTs and gold such as large surface area and high electrochemical conductivity, as well as high affinity and selectivity of aptamer toward its target and the stronger interaction of SWNTs with single-stranded DNA (ssDNA) than double-stranded DNA (dsDNA). In the absence of cocaine, a little amount of SWNTs bind to Aptamer-CS-modified electrode, so that the electrochemical signal is weak. In the presence of cocaine, aptamer binds to cocaine, leaves the surface of electrode. So that, a large amount of SWNTs bind to CS-modified electrode, generating to a strong electrochemical signal. The designed electrochemical aptasensor showed good selectivity toward cocaine with a limit of detection (LOD) as low as 105 pM. Moreover, the fabricated electrochemical aptasensor was successfully applied to detect cocaine in serum with a LOD as low as 136 pM.

Ultrasensitive detection of lead (II) based on fluorescent aptamer-functionalized carbon nanotubes.

Lead contamination is a serious environmental problem with toxic effects in human. Here, we developed a simple and sensitive sensing method employing ATTO 647N/aptamer-SWNT ensemble for detection of Pb(2+). This method is based on the super quenching capability of single-walled carbon nanotubes (SWNTs), high affinity of the aptamer toward Pb(2+) and different propensities of ATTO 647N-aptamer and ATTO 647N-aptamer/Pb(2+) complex for adsorption on SWNTs. In the absence of Pb(2+), the fluorescence of ATTO 647N-aptamer is efficiently quenched by SWNTs. Upon addition of Pb(2+), the aptamer binds to its target, leading to the formation of a G-quadruplex/Pb(2+) complex and does not interact with SWNTs and ATTO 647N-aptamer starts fluorescing. This sensor exhibited a high selectivity toward Pb(2+) and a limit of detection (LOD) as low as 0.42 nM was obtained. Also this sensor could be applied for detection of Pb(2+) ions in tap water and biological sample like serum with high sensitivity.