RESUMEN
An electrochemical aptamer-based sensor was developed for glutamate, the major excitatory neurotransmitter in the central nervous system. Determining glutamic acid release and glutamic acid levels is crucial for studying signal transmission and for diagnosing pathological conditions in the brain. Glutamic acid-selective oligonucleotides were isolated from an ssDNA library using the Capture-SELEX protocol in complex medium. The selection permitted the isolation of an aptamer 1d04 with a dissociation constant of 12 µM. The aptamer sequence was further used in the development of an electrochemical aptamer sensor. For this purpose, a truncated aptamer sequence named glu1 was labelled with a ferrocene redox tag at the 3'-end and immobilized on a gold electrode surface via Au-thiol bonds. Using 6-mercapto-1-hexanol as the backfill, the sensor performance was characterized by alternating current voltammetry. The glu1 aptasensor showed a limit of detection of 0.0013 pM, a wide detection range between 0.01 pM and 1 nM, and good selectivity for glutamate in tenfold diluted human serum. With this enzyme-free aptasensor, the highly selective and sensitive detection of glutamate was demonstrated, which possesses great potential for implementation in microelectrodes and for in vitro as well as in vivo monitoring of neurotransmitter release.
Asunto(s)
Aptámeros de Nucleótidos/química , Técnicas Electroquímicas/métodos , Ácido Glutámico/sangre , Técnicas Biosensibles/métodos , Ácido Glutámico/análisis , Hexanoles/química , Humanos , Límite de Detección , Compuestos de Sulfhidrilo/químicaRESUMEN
In this present work, we proposed a colorimetric strategy for simultaneous detection of histidine and cysteine based on G-quadruplex-Cu(II) metalloenzyme for the first time. Because of the adding of histidine or cysteine, the formation of G-quadruplex-Cu(II) metalloenzyme will be disturbed, thus the catalytic activity to TMB-H2O2 reaction is inversely proportional to the concentration of histidine or cysteine. With this strategy, the limit of detection in experimental measurement for histidine and cysteine is 10 nM and 5 nM, respectively, which are both lower than previous colorimetric arrays. With the help of NEM, cysteine is alkylated and the reaction between Cu(2+) is inhibited, so the selectivity can also be guaranteed. The cost is quite low since the developed array is label free and enzyme free by using low-cost DNA and Cu(2+). More importantly, the colorimetric detection operation is very simple without any further modification process.
Asunto(s)
Colorimetría/métodos , Cisteína/análisis , Enzimas/química , G-Cuádruplex , Histidina/análisis , Metaloproteínas/química , Límite de DetecciónRESUMEN
Multielectrode arrays (MEAs) have been increasingly used for the development of biosensors due to their capability to record signals from multiple channels, fast mass transfer rates, and high spatial resolution. Alzheimer's disease (AD) is often associated with mitochondrial dysfunction, which is closely related to reduced levels of adenosine triphosphate (ATP). Therefore, simultaneous detection of ATP together with amyloid-ß oligomers (AßO), a reliable biomarker for AD, can potentially advance the early detection of Alzheimer's disease. In this work, a dual-aptamer modified MEA chip was developed that consists of microelectrodes modified with electrodeposited 3D nanostructures (3D-GMEs). Electrodeposition methods, deposition potential, and deposition time were systematically altered and the active surface areas as well as the electrode morphologies were characterized by cyclic voltammetry and scanning electron microscopy. The nanostructured microelectrodes were sequentially modified with AßO and ATP specific aptamer receptors. To achieve the modification of different aptamer receptors at different 3D-GMEs of the same MEA chip, electrochemical cleaning was applied to individual 3D-GMEs. Ferrocene labels were attached to the aptamer receptors to enable amperometric signaling after target-aptamer binding. The developed aptasensor showed a linear detection range from 1 pM to 200 nM for the detection of AßO and from 0.01 nM to 1000 nM for the detection of ATP. Finally, ATP and AßO were detected simultaneously in the same analyte solution by the same sensor chip, which could support the early detection of AD, provide comprehensive information about the health status of the patient, and be helpful for pathological studies of neurodegenerative diseases.
Asunto(s)
Adenosina Trifosfato/líquido cefalorraquídeo , Péptidos beta-Amiloides/líquido cefalorraquídeo , Aptámeros de Nucleótidos/química , Técnicas Biosensibles , Enfermedad de Alzheimer/diagnóstico , Biomarcadores/líquido cefalorraquídeo , Técnicas Electroquímicas , Oro/química , Humanos , Límite de Detección , Microelectrodos , Nanoestructuras/químicaRESUMEN
Better approaches are critically needed for in situ point-of-care diagnostic biosensors that enable primary care physicians, or even individual patients, to directly analyze biological fluids without complicated sample pretreatments. Additional purification steps consume time, consume reagents, often require other equipment, and can introduce false-negative results. Biosensors have been modified with blocking molecules to reduce biofouling; however, the effectiveness relies on their chemical composition and morphology. Here, we used a polyethylene glycol film to suppress unspecific binding from human serum on an electrochemical malaria aptasensor. A detailed study of the variation of the chemical and morphological composition of the aptamer/polyethylene glycol mixed monolayer as a function of incubation time was conducted. Higher resistance to matrix biofouling was found for polyethylene glycol than for hydrophobic alkanethiol films. The best sensor performance was observed for intermediate polyethylene glycol immobilization times. With prolonged incubation, phase separation of aptamer, and polyethylene glycol molecules locally increased the aptamer density and thereby diminished the analyte binding capability. Remarkably, polyethylene glycols do not affect the aptasensor sensitivity but enhance the complex matrix tolerance, the dynamic range, and the limit of detection. Careful tuning of the blocking molecule immobilization is crucial to achieving high aptasensor performance and biofouling resistance.
Asunto(s)
Aptámeros de Nucleótidos/química , Técnicas Electroquímicas/instrumentación , Malaria/diagnóstico , Polietilenglicoles/química , Biomarcadores/sangre , Humanos , L-Lactato Deshidrogenasa/metabolismo , Límite de Detección , Microscopía de Fuerza Atómica , Plasmodium falciparum/enzimologíaRESUMEN
Electrochemical aptamer receptor/transducer systems are key elements of emerging E-AB sensors (aptasensor) used for the detection of various kinds of targets. However, the performance of these amperometric sensors is often limited by the low density of receptors attached to the sensor surface and high background signals. In the present work, interdigitated organic electrochemical transistors (iOECT) were used as a transducer to enhance the sensitivity and dynamic detection range of aptasensors. Therefore, the electrode of an amperometric sensor was utilized as gate electrode to operate the iOECT. This device was used to detect the low weight target molecule adenosine triphosphate (ATP), a common biomarker, which plays an important role for cardiovascular, neurodegenerative, and immune deficiency diseases. The novel aptasensor can selectively detect ATP with ultrahigh sensitivity down to the concentration of 10 pM, which is four orders of magnitude lower than the detection limit of the same aptasensor using an amperometric transducer principle (limit-of-detection of 106â¯nM) and most other previously reported electrochemical sensors. Furthermore, sensor regeneration was demonstrated, which facilitates reusability of OECT aptasensors. The small device size in combination with high transconductances paves the way for the development of highly sensitive integrated micro-biosensors for point-of-care applications.
Asunto(s)
Adenosina Trifosfato/aislamiento & purificación , Aptámeros de Nucleótidos/química , Técnicas Biosensibles , Técnicas Electroquímicas , Adenosina Trifosfato/química , Electrodos , Oro , Humanos , Límite de DetecciónRESUMEN
For the first time by integrating fluorescent polyT-templated CuNPs and SYBR Green I, a basic INHIBIT gate and four advanced logic circuits (2-to-1 encoder, 4-to-2 encoder, 1-to-2 decoder and 1-to-2 demultiplexer) have been conceptually realized under label-free and enzyme-free conditions. Taking advantage of the selective formation of CuNPs on ss-DNA, the implementation of these advanced logic devices were achieved without any usage of dye quenching groups or other nanomaterials like graphene oxide or AuNPs since polyA strands not only worked as an input but also acted as effective inhibitors towards polyT templates, meeting the aim of developing bio-computing with cost-effective and operationally simple methods. In short, polyT-templated CuNPs, as promising fluorescent signal reporters, are successfully applied to fabricate advanced logic devices, which may present a potential path for future development of molecular computations.
RESUMEN
Multiple advanced logic circuits including the full-adder, full-subtract and majority logic gate have been successfully realized on a DNA/GO platform for the first time. All the logic gates were implemented in an enzyme-free condition. The investigation provides a wider field of vision towards prototypical DNA-based algebra logical operations and promotes the development of advanced logic circuits.
Asunto(s)
Computadores Moleculares , ADN , Grafito , Lógica , ÓxidosRESUMEN
In this work, we have successfully realized multivalued logic circuits including ternary INHIBIT and ternary OR logic gates in an enzyme-free condition by integration of graphene oxide and DNA for the first time. Compared to the binary logic gate with two states of "0" and "1", the multivalued logic gate contains three different states of "0", "1", and "2", which can increase the information content in a system and further improve the ability of information processing. Such types of multivalued logic operations provide a wider field of vision toward DNA-based algebra logical operations to make applications more accurate with complexity reduction and accelerate the development of advanced logic gates.
RESUMEN
With CCRF-CEM as the model cell, a highly sensitive and selective cytosensor was developed by taking advantage of polydopamine nanospheres for the first time. The strategies of aptamer/membrane protein recognition and Exonuclease III assisted cycle amplification were used for improving selectivity and sensitivity. The detection of limit reached was as low as 15 cells per mL.
Asunto(s)
Aptámeros de Nucleótidos/química , Técnicas Biosensibles , Exodesoxirribonucleasas/química , Indoles/química , Nanosferas/química , Polímeros/química , Animales , Bovinos , Moléculas de Adhesión Celular/química , Línea Celular Tumoral , Transferencia Resonante de Energía de Fluorescencia , Humanos , Límite de Detección , Tamaño de la Partícula , Proteínas Tirosina Quinasas Receptoras/química , Albúmina Sérica Bovina/químicaRESUMEN
A DNA-based 2:1 multiplexer and 1:2 demultiplexer have been conceptually realized in enzyme-free conditions. For the first time, the designed DNA-based multiplexer could be implemented by keeping input/output signal homogeneity, which has great potential application in information processing.
Asunto(s)
Computadores Moleculares , ADN de Cadena Simple/química , Grafito/química , Óxidos/química , Fluoresceínas/química , Colorantes Fluorescentes/química , G-Cuádruplex , Mesoporfirinas/química , Hibridación de Ácido NucleicoRESUMEN
A logically reversible Feynman gate was successfully realized under enzyme-free conditions by integrating graphene oxide and DNA for the first time. The gate has a one-to-one mapping function to identify inputs from the corresponding outputs. This type of reversible logic gate may have great potential applications in information processing and biosensing systems.