G-Quadruplex-based DNAzyme for colorimetric detection of cocaine: using magnetic nanoparticles as the separation and amplification element

The appearance of the aptamer provides good recognition elements for small molecules, especially for drugs. In this work, by combining the advantages of magnetic nanoparticles (MNPs) with colorimetric drug detection using hemin-G-quadruplex complex as the sensing element, we report a simple and sensitive DNAzyme-based colorimetric sensor for cocaine detection in a 3,3,5,5-tetramethylbenzidine sulfate (TMB)-H(2)O(2) reaction system. The whole experimental processes are simplified. Cocaine aptamer fragments, SH-C2, are covalently labeled onto the amine-functionalized MNPs. When the target cocaine and another cocaine aptamer fragments (C1) grafted with G-riched strand AG4 (i.e. C1-AG4) are present simultaneously, the C2 layer on MNPs hybridizes partly with C1-AG4 to bind the cocaine. The C1-AG4 can be combinded with hemin to form DNAzyme which can effectively catalyze the H(2)O(2)-mediated oxidation of TMB, giving rise to a change in solution color. Importantly, using MNPs as the separation and amplification elements could effectively reduce the background signal and the interference from the real samples. A linear response from 0.1 μM to 20 μM is obtained for cocaine and a detection limit of 50 nM is achieved, which provides high sensitivity and selectivity to detect cocaine.     

Aptamer sandwich-based carbon nanotube sensors for single-carbon-atomic-resolution detection of non-polar small molecular species

A portable sensor platform for the detection of small molecular species is crucial for the on-site monitoring of environmental pollutants, food toxicants, and disease-related metabolites. However, it is still extremely difficult to find highly selective and sensitive sensor platforms for general small molecular detection. Herein, we report aptamer sandwich-based carbon nanotube sensor strategy for small molecular detection, where aptamers were utilized to capture target molecules as well as to enhance the sensor signals. We successfully demonstrated the detection of non-polar bisphenol A molecules with a 1 pM sensitivity. Significantly, our sensors were able to distinguish between similar small molecular species with single-carbon-atomic resolution. Furthermore, using the additional biotin modification on labeling aptamer, we enhanced the detection limit of our sensors down to 10 fM. This strategy allowed us to detect non-polar small molecular species using carbon nanotube transistors, thus overcoming the fundamental limitation of field effect transistor-based sensors. Considering the extensive applications of sandwich assay for the detection of rather large biomolecules, our results should open up completely new dimension in small molecular detection technology and should enable a broad range of applications such as environmental protection and food safety.     

Optimization of electrochemical aptamer-based sensors via optimization of probe packing density and surface chemistry

Electrochemical, aptamer-based (E-AB) sensors, which are comprised of an electrode modified with surface immobilized, redox-tagged DNA aptamers, have emerged as a promising new biosensor platform. In order to further improve this technology we have systematically studied the effects of probe (aptamer) packing density, the AC frequency used to interrogate the sensor, and the nature of the self-assembled monolayer (SAM) used to passivate the electrode on the performance of representative E-AB sensors directed against the small molecule cocaine and the protein thrombin. We find that, by controlling the concentration of aptamer employed during sensor fabrication, we can control the density of probe DNA molecules on the electrode surface over an order of magnitude range. Over this range, the gain of the cocaine sensor varies from 60% to 200%, with maximum gain observed near the lowest probe densities. In contrast, over a similar range, the signal change of the thrombin sensor varies from 16% to 42% and optimal signaling is observed at intermediate densities. Above cut-offs at low hertz frequencies, neither sensor displays any significant dependence on the frequency of the alternating potential employed in their interrogation. Finally, we find that E-AB signal gain is sensitive to the nature of the alkanethiol SAM employed to passivate the interrogating electrode; while thinner SAMs lead to higher absolute sensor currents, reducing the length of the SAM from 6-carbons to 2-carbons reduces the observed signal gain of our cocaine sensor 10-fold. We demonstrate that fabrication and operational parameters can be varied to achieve optimal sensor performance and that these can serve as a basic outline for future sensor fabrication.     

Label-free and sensitive impedimetric nanosensor for the detection of cocaine based on a supramolecular complexation with β-cyclodextrin,immobilized on a nanostructured polymer film

Cocaine, a powerful addictive stimulant drug, has a variety of adverse effects on the body, thus its sensitive detection is very important. Here, we report on a simple, label-free, and sensitive impedimetric sensor for determination of cocaine based on its affinity to form an inclusion complex with β-cyclodextrin (β-CD). First, we prepared nanostructured poly N-acetylaniline film via electropolymerization of its monomer on a glassy carbon electrode (PNAANI/GC), subsequently overoxidized it, and conjugated β-CD to the polymer backbone. The designed and synthesized nanostructured PNAANI film serves a dual function in the sensor: on one hand, it maintains a high effective surface area on a geometrically small electrode that significantly enhances the number of β-CD molecules immobilized on the electrode; on the other hand, it provides an upright-oriented β-CD conjugation to the polymer backbone, thus all the β-CD receptors are actively involved in responding to the target. Sensitivity of the sensor was further enhanced by preconcentration of cocaine on the modified electrode surface. We attributed the changes in the interfacial charge transfer resistance (R _ct) of the electrode to cocaine concentration. Under optimized condition (pH 7.4, 5-min accumulation at an open circuit voltage), the sensor responded to cocaine concentration in the range of 100 nM–1.0 mM with a detection limit of 50 nM. Selectivity of the sensor for cocaine relative to some potential inferring compounds was also investigated, and the results were promising. The proposed approach exhibited an extended dynamic range, low detection limit, good sensitivity, and a desirable selectivity, which provides an efficient application prospect for on-field cocaine sensing.     

Design of Turn‐On Near‐Infrared Fluorescent Probes for Highly Sensitive and Selective Monitoring of Biopolymers

Simple, sensitive, and selective detection of specific biopolymers is critical in a broad range of biomedical and technological areas. We present a design of turn‐on near‐infrared (NIR) fluorescent probes with intrinsically high signal‐to‐background ratio. The fluorescent signal generation mechanism is based on the aggregation/de‐aggregation of phthalocyanine chromophores controlled by selective binding of small‐molecule “anchor” groups to a specific binding site of a target biopolymer. As a proof‐of‐concept, we demonstrate a design of a sensor for EGFR tyrosine kinase—an important target in cancer research. The universality of the fluorescent signal generation mechanism, as well as the dependence of the response selectivity on the choice of the small‐molecule “anchor” group, make it possible to use this approach to design reliable turn‐on NIR fluorescent sensors for detecting specific protein targets present in the low‐nanomolar concentration range.     

Sensor Array: Impedimetric Label‐Free Sensing of DNA Hybridization in Real Time for Rapid,PCR‐Based Detection of Microorganisms

A novel, impedance‐based electronic sensor format was used for label‐free, real‐time detection of microbial DNA on oligonucleotide probe arrays. Detection limits of 5–10 nM were achieved for single‐stranded, PCR‐amplified DNA targets. Hybridization selectivity yielded 9‐ to 12‐fold signal increases for specific targets, and the sensor arrays were re‐used multiple times without significant signal degradation. These and other features of the SHARP Laboratories of America (SLA) sensor array, such as its ability to acquire continuous measurements of DNA as it accumulates on the array surface, make it an attractive biosensor platform for field detection and monitoring of sentinel and/or pathogenic microorganisms.     

Conjugated polyelectrolyte amplified fluorescent assays with probe functionalized silica nanoparticles for chemical and biological sensing

Low-cost sensors with high sensitivity and selectivity for chemical and biological detection are of high scientific and economic importance. Silica nanoparticles (NPs) have shown vast promise in sensor applications by virtue of their controllable surface modification, good chemical stability, and biocompatibility. This mini-review summarizes our recent development of silica NP-based assays for chemical and biological detection, where silica NPs serve as the substrate for probe immobilization, target recognition, and separation. The assay performance is further improved through the introduction of conjugated polyelectrolyte to amplify the detection signal. The assays have been demonstrated to be successful for the detection of DNA, small molecules, and proteins. They could be generalized for other targets based on specific interactions, such as DNA hybridization, antibody-antigen recognition, and target-aptamer binding.     

Interactions of DNA with graphene and sensing applications of graphene field-effect transistor devices: A review

Graphene field-effect transistors (GFET) have emerged as powerful detection platforms enabled by the advent of chemical vapor deposition (CVD) production of the unique atomically thin 2D material on a large scale. DNA aptamers, short target-specific oligonucleotides, are excellent sensor moieties for GFETs due to their strong affinity to graphene, relatively short chain-length, selectivity, and a high degree of analyte variability. However, the interaction between DNA and graphene is not fully understood, leading to questions about the structure of surface-bound DNA, including the morphology of DNA nanostructures and the nature of the electronic response seen from analyte binding. This review critically evaluates recent insights into the nature of the DNA graphene interaction and its affect on sensor viability for DNA, small molecules, and proteins with respect to previously established sensing methods. We first discuss the sorption of DNA to graphene to introduce the interactions and forces acting in DNA based GFET devices and how these forces can potentially affect the performance of increasingly popular DNA aptamers and even future DNA nanostructures as sensor substrates. Next, we discuss the novel use of GFETs to detect DNA and the underlying electronic phenomena that are typically used as benchmarks for characterizing the analyte response of these devices. Finally, we address the use of DNA aptamers to increase the selectivity of GFET sensors for small molecules and proteins and compare them with other, state of the art, detection methods.     

Chemical vapor detection using single-walled carbon nanotubes

Single-walled carbon nanotubes possess unique properties that make them a potentially ideal material for chemical sensing. However, their extremely small size also presents technical challenges for realizing a practical sensor technology. In this tutorial review we explore the transduction physics by which the presence of molecular adsorbates is converted into a measurable electronic signal, and we identify solutions to the problems such as nanotube device fabrication and large, low-frequency noise that have inhibited commercial sensor development. Finally, we examine strategies to provide the necessary chemical specificity to realize a nanotube-based detection system for trace-level chemical vapor detection.     

Universal quantum dot-based sandwich-like immunoassay strategy for rapid and ultrasensitive detection of small molecules using portable and reusable optofluidic nano-biosensing platform

A universal sandwich-like immunoassay strategy based on quantum-dots immunoprobe (QD-labeled anti-mouse IgG antibody) was developed for rapid and ultrasensitive detection of small molecules. A portable and reusable optofluidic nano-biosensing platform was applied to investigate the sandwich-like immunoassay mechanism and format of small molecules, as well as the binding kinetics between QD immunoprobe and anti-small molecule antibody. A two-step immunoassay method that involves pre-incubation mixture of different concentration of small molecule and anti-small molecule antibody, and subsequent introduction of QD immunoprobe into the optofluidic cell was conducted for small molecule determination. Compared with the one-step immunoassay method, the two-step immunoassay method can obtain higher fluorescence signal and higher sensitivity index, thus improving the nano-biosensing performance. Based on the proposed strategy, two mode targets, namely, microcystin-LR (MC-LR) and Bisphenol A (BPA) were tested with high sensitivity, rapidity, and ease of use. A higher concentration of small molecules in the sample led to less anti-small molecule antibody bound with antigen-carrier protein conjugate immobilized onto the sensor surface, and less QD immunoprobes bound with anti-small molecule antibody. This phenomenon lowered the fluorescence signal detected by nano-biosensing platform. Under optimal operating conditions, MC-LR and BPA exhibited a limit of detection of 0.003 and 0.04 μg/L, respectively. The LODs were better than those of the indirect competitive immunoassay method for small molecules via Cy5.5-labeled anti-small molecule antibody. The proposed QD-based sandwich-like immunoassay strategy was evaluated in spiked water samples, and showed good recovery, precision and accuracy without complicated sample pretreatments. All these results demonstrate that the new detection strategy could be readily applied to the other trace small molecules in real water samples.     

State of the art: Lateral flow assay (LFA) biosensor for on-site rapid detection

We summarized the principle of LFAs, three main elements for the LFAs (recognition molecule, signal transduction element, the targets) and different optimal experimental conditions in the recent LFA-related studies to give detailed overview of the LFA development.     

Non-labeling multiplex surface enhanced Raman scattering (SERS) detection of volatile organic compounds (VOCs)

In this paper, we report multiplex SERS based VOCs detection with a leaning nano-pillar substrate. The VOCs analyte molecules adsorbed at the tips of the nano-pillars produced SERS signal due to the field enhancement occurring at the localized surface plasmon hot spots between adjacent leaning nano-pillars. In this experiment, detections of acetone and ethanol vapor at different concentrations were demonstrated. The detection limits were found to be 0.0017 ng and 0.0037 ng for ethanol and acetone vapor molecules respectively. Our approach is a non-labeling method such that it does not require the incorporation of any chemical sensing layer for the enrichment of gas molecules on sensor surface. The leaning nano-pillar substrate also showed highly reproducible SERS signal in cyclic VOCs detection, which can reduce the detection cost in practical applications. Further, multiplex SERS detection on different combination of acetone and ethanol vapor was also successfully demonstrated. The vibrational fingerprints of molecular structures provide specific Raman peaks for different VOCs contents. To the best of our knowledge, this is the first multiplex VOCs detection using SERS. We believe that this work may lead to a portable device for multiplex, specific and highly sensitive detection of complex VOCs samples that can find potential applications in exhaled breath analysis, hazardous gas analysis, homeland security and environmental monitoring.     

Aptamer/quantum dot-based simultaneous electrochemical detection of multiple small molecules

A novel strategy for “signal on” and sensitive one-spot simultaneous detection of multiple small molecular analytes based on electrochemically encoded barcode quantum dot (QD) tags is described. The target analytes, adenosine triphosphate (ATP) and cocaine, respectively, are sandwiched between the corresponding set of surface-immobilized primary binding aptamers and the secondary binding aptamer/QD bioconjugates. The captured QDs yield distinct electrochemical signatures after acid dissolution, whose position and size reflect the identity and level, respectively, of the corresponding target analytes. Due to the inherent amplification feature of the QD labels and the “signal on” detection scheme, as well as the sensitive monitoring of the metal ions released upon acid dissolution of the QD labels, low detection limits of 30 nM and 50 nM were obtained for ATP and cocaine, respectively, in our assays. Our multi-analyte sensing system also shows high specificity to target analytes and promising applicability to complex sample matrix, which makes the proposed assay protocol an attractive route for screening of small molecules in clinical diagnosis.     

Design of Turn-On Near-Infrared Fluorescent Probes for Highly Sensitive and Selective Monitoring of Biopolymers

Simple, sensitive, and selective detection of specific biopolymers is critical in a broad range of biomedical and technological areas. We present a design of turn-on near-infrared (NIR) fluorescent probes with intrinsically high signal-to-background ratio. The fluorescent signal generation mechanism is based on the aggregation/de-aggregation of phthalocyanine chromophores controlled by selective binding of small-molecule “anchor” groups to a specific binding site of a target biopolymer. As a proof-of-concept, we demonstrate a design of a sensor for EGFR tyrosine kinase—an important target in cancer research. The universality of the fluorescent signal generation mechanism, as well as the dependence of the response selectivity on the choice of the small-molecule “anchor” group, make it possible to use this approach to design reliable turn-on NIR fluorescent sensors for detecting specific protein targets present in the low-nanomolar concentration range.     

Direct detection of low-concentration NO in physiological solutions by a new GaAs-based sensor

Nitric oxide (NO) acts as a signal molecule in the nervous system, as a defense against infections, as a regulator of blood pressure, and as a gate keeper of blood flow to different organs. In vivo, it is thought to have a lifetime of a few seconds. Therefore, its direct detection at low concentrations is difficult. We report on a new type of hybrid, organic-semiconductor, electronic sensor that makes detection of nitric oxide in physiological solution possible. The mode of action of the device is described to explain how its electrical resistivity changes as a result of NO binding to a layer of native hemin molecules. These molecules are self-assembled on a GaAs surface to which they are attached through a carboxylate binding group. The new sensor provides a fast and simple method for directly detecting NO at concentrations down to 1 microM in physiological aqueous (pH=7.4) solution at room temperature.     

Application of split aptamers-based sensors for the detection of small moleculesEI北大核心CSCD

The split aptamers (SPAs)-based sensors is a novel kind of biosensors, assembled by two or more oligonucleotides in the presence of specific targets. To be successfully assembled, the sensor has to be induced by a specific target, which can aviod false-positive results and thus has a high degree of specificity and sensitivity. SPAs are suitable for the detection of various targets and show great advantages and potential in the development of aptasensors, especially for the detection of small molecules. However, the development and application of SPA-based sensors still remain challenging. Currently, the major difficulty is how to improve the stability of SPA-target complexes. Herein, this review summarizes the SPAs, strategies of splitting aptamers, and their applications in the detection of small molecules, aiming to provide new ideas for the development of novel, sensitive, and specific aptasensors. © 2023, Youke Publishing Co.,Ltd. All rights reserved.     

基于多边形金纳米颗粒的非标记型可卡因适体传感器的研制

实验合成了多边形金纳米颗粒,通过壳聚糖(CHIT)将合成的多边形金纳米颗粒固定在玻碳电极表面,然后通过自组装技术将带巯基的捕获DNA探针固定在修饰有多边形金纳米颗粒的电极表面,利用杂交反应使可卡因适体与DNA捕获探针结合,制成非标记型可卡因适体传感器。以六氨合钌作为电化学指示剂,通过测量传感器与目标物可卡因结合前后电流变化情况对可卡因进行测定。考察了缓冲溶液的pH、可卡因培育时间、扫描速度等对测定的影响。结果表明,在pH为7.40时该传感器的检测范围为1.0×10-10～1.0×10-3 mol/L,检测限为3.0×10-11 mol/L。该传感器制作简单,响应好,抗干扰能力强。     

Aptamer functionalized microcantilever sensors for cocaine detection

A cocaine-specific aptamer was used as a receptor molecule in a microcantilever-based surface stress sensor for detection of cocaine molecules. An interferometric technique that relies on measuring differential displacement between two microcantilevers (a sensing/reference pair) was utilized to measure the cocaine/aptamer binding induced surface stress changes. Sensing experiments were performed for different concentrations of cocaine from 25 to 500 μM in order to determine the sensor response as a function of cocaine concentration. In the lower concentration range from 25 to 100 μM, surface stress values increased proportionally to coverage of aptamer/cocaine complexes from 11 to 26 mN/m. However, as the cocaine concentration was increased beyond 100 μM, the surface stress values demonstrated a weaker dependence on the affinity complex surface coverage. On the basis of a sensitivity of 3 mN/m for the surface stress measurement, the lowest detectable threshold for the cocaine concentration is estimated to be 5 μM. Sensing cantilevers could be regenerated and reused because of reversible thermal denaturation of aptamer.     

Label‐free Electrochemiluminescent Enantioselective Sensor for Distinguishing between Chiral Metallosupramolecular Complexes

Chiral molecular recognition of DNA is important for rational drug design and for developing structural probes of DNA conformation. Developing a convenient and inexpensive assay for sensitive and selective identification of DNA‐specific binding compounds with rapid, easy manipulation is in ever‐increasing demand. Here, we present a “turn‐on” and label‐free electrochemiluminescent (ECL) biosensor for distinguishing chiral metallosupramolecular complexes based on DNA three‐way junction formation selectively induced by the analyte. The fabricated ECL sensor shows excellent performance in the chiral discrimination of two enantiomers with an enantioselective recognition ratio of up to 4.4. More importantly, as a “turn‐on” detection system, the ECL chiral sensor does not suffer from false positives and limited signal range of “signal‐off” systems. Therefore, this concept may provide a new insight into the design of efficient sensors for distinguishing chiral molecules and for investigating the interactions between DNA and small molecules.     

DNA detection method based on the two-dimensional aggregation and selective desorption of nanoparticle probes

A label-free two-dimensional colorimetric DNA sensor is reported. This sensor is based on the 2D aggregation of oligonucleotide-modified gold nanoparticle probes induced by the molecular hybridization of single-stranded oligonucleotide probes and their complementary single-stranded DNA targets. To detect the aggregation, we have developed a new detection method based on the selective desorption of nonaggregated nanoparticles. We will show here that this detection method is highly specific and allows the quantification of the DNA targets.