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A Label-Free Fluorescent DNA Calculator Based on Gold Nanoparticles for Sensitive Detection of ATP
摘要： Herein we described a deoxyribonucleic acid (DNA) calculator for sensitive detection of the determination of adenosine triphosphate (ATP) using gold nanoparticles (GNP) and PicoGreen fluorescence dye as signal transducer, and ATP and single-stranded DNA (DNA-M') as activators. The calculator-related performances including linearity, reaction time, logic gate, and selectivity were investigated, respectively. The results revealed that this oligonucleotide sensor was highly sensitive and selective. The detection range was 50–500 nmol/L (R2 = 0.99391) and the detection limit was 46.5 nmol/L. The AND DNA calculator was successfully used for the ATP detection in human urine. Compared with other methods, this DNA calculator has the characteristics of being label-free, non-enzymic, simple, and highly sensitive.
关键词： DNA calculator,enzyme-free,gold nanoparticles,label-free fluorescence,ATP detection
更新于2025-11-19 16:46:39
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A Label-Free Fluorescent DNA Machine for Sensitive Cyclic Amplification Detection of ATP
摘要： In this study, a target recycled ampli?cation, background signal suppression, label-free ?uorescent, enzyme-free deoxyribonucleic acid (DNA) machine was developed for the detection of adenosine triphosphate (ATP) in human urine. ATP and DNA fuel strands (FS) were found to trigger the operation of the DNA machine and lead to the cyclic multiplexing of ATP and the release of single stranded (SS) DNA. Double-stranded DNA (dsDNA) was formed on graphene oxide (GO) from the combination of SS DNA and complementary strands (CS(cid:48)). These double strands then detached from the surface of the GO and in the process interacted with PicoGreen dye resulting in amplifying ?uorescence intensity. The results revealed that the detection range of the DNA machine is from 100 to 600 nM (R2 = 0.99108) with a limit of detection (LOD) of 127.9 pM. A DNA machine circuit and AND-NOT-AND-OR logic gates were successfully constructed, and the strategy was used to detect ATP in human urine. With the advantage of target recycling ampli?cation and GO suppressing background signal without ?uorescent label and enzyme, this developed strategy has great potential for sensitive detection of different proteins and small molecules.
关键词： cyclic ampli?cation,ATP detection,DNA machine,label-free ?uorescence,graphene oxide,logic gate
更新于2025-11-19 16:46:39
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Graphene foam field-effect transistor for ultra-sensitive label-free detection of ATP
摘要： As the major energy molecule of cells, adenosine triphosphate (ATP) regulates various biological processes and has been found to be closely related to many diseases. Therefore, ATP detection in trace amounts is very useful for understanding various biological processes, studying cellular events such as proliferation and apoptosis, and estimating contaminated degree of food and medical instrument. To date, the trace sensing ATP at picomolar level in biological systems is still a major challenge. Because of unique electrical and structural properties, graphene has attracted much attention in biosensing applications. Here, a sensitive and selective graphene foam field-effect transistor (GF-FET) biosensor for ATP detection is demonstrated. The lowest detection limit of the biosensors for analyzing ATP is down to 0.5 pM, which is one or several orders lower than the reported results. Moreover, the GF-FET biosensor show a good linear current response to ATP concentrations in a broad range from 0.5 pM to 50 μM. The GF-FET sensor surface can be regenerated for many times and used for up to weeks without significant loss of functionality. Based on this sensing platform, label-free measurements of ATP concentrations in human serum as well as in cell lysate are demonstrated. The work may provide a novel platform to study ATP release and energy-regulated biological processes, suggesting a promising future for biosensing applications.
关键词： FET,ATP,Graphene foams
更新于2025-09-23 15:23:52
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A fluorometric method for determination of the activity of T4 polynucleotide kinase by using a DNA-templated silver nanocluster probe
摘要： The authors describe a turn-off fluorometric method for the determination of the activity of the T4 polynucleotide kinase (T4 PNK). It is based on the use of DNA-templated silver nanoclusters (AgNCs). DNA probes with terminal 5′ hydroxy groups are used as substrates for DNA phosphatases. If subsequently treated with T4 PNK and Lambda exonuclease (λ exo), the AgNC DNA probes with a modified C-rich sequence and the G-rich sequence is separated. Upon their separation, the strong fluorescence (with excitation/emission maxima at 580/650 nm) that is caused by the proximity of the G-rich region and the C-rich region in the AgNCs decreases sharply. This enabled the fluorometric kinetic determination of the activity of T4 PNK. The assay is characterized by a wide linear range (from 0.01 to 12.5 U·mL?1), a low detection limit (0.01 U·mL?1) and short assay time (typically 60 min). This makes it a promising tool for use in studying processes related to DNA phosphorylation, in drug discovery and in diagnostics.
关键词： DNA-AgNCs,Cell extracts,Clinical diagnostics,Lambda exonuclease,Proximity effect,T4 PNK,Inhibitor,ATP,Na2HPO4,Phosphorylation
更新于2025-09-23 15:23:52
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Ultrasensitive detection of avian influenza A (H7N9) virus using surface-enhanced Raman scattering-based lateral flow immunoassay strips
摘要： The development of biosensors that are portable, low-cost, and quantitative has long been sought for rapid, on-site, and timely detection of avian influenza virus (AIV). In this study, an antibody-based Raman lateral flow immunoassay strip was developed to detect AIV H7N9. This LFIA strip used a novel core-shell structure material, AuAg4(cid:3)ATP@AgNPs, as a Raman probe. An antibody specific for AIV and goat anti-mouse IgG antibody were immobilized on a nitrocellulose membrane as the test and control lines, respectively. Accumulation of antibody-virus-antibody-Raman probe complex at the test line could be visualized by the naked eye, and the Raman signal could be quantified using a portable Raman instrument. The testing process for the SERS-based LFIA strips could be completed in 20 min, which avoided the time-cost of current methods for AIV analysis. In our SERS-based biosensor, we estimated the limit of detection (LOD) for H7N9 to be 0.0018 HAU. This value is approximately three orders of magnitude more sensitive than the corresponding HA assays. When testing real sample, the results of the strip test were in accordance with those from real-time PCR testing. In conclusion, the SERS-based LFIA strip proposed in this study shows tremendous potential to detect targets quickly and sensitively using an elegantly simple method.
关键词： AuAg4(cid:3)ATP@AgNPs,Surface-enhanced Raman scattering,Avian influenza virus,Lateral flow immunoassay strips
更新于2025-09-23 15:23:52
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Step Sizes and Rate Constants of Single-headed Cytoplasmic Dynein Measured with Optical Tweezers
摘要： A power stroke of dynein is thought to be responsible for the stepping of dimeric dynein. However, the actual size of the displacement driven by a power stroke has not been directly measured. Here, the displacements of single-headed cytoplasmic dynein were measured by optical tweezers. The mean displacement of dynein interacting with microtubule was ~8 nm at 100 μM ATP, and decreased sigmoidally with a decrease in the ATP concentration. The ATP dependence of the mean displacement was explained by a model that some dynein molecules bind to microtubule in pre-stroke conformation and generate 8-nm displacement, while others bind in the post-stroke one and detach without producing a power stroke. Biochemical assays showed that the binding affinity of the post-stroke dynein to a microtubule was ~5 times higher than that of pre-stroke dynein, and the dissociation rate was ~4 times lower. Taking account of these rates, we conclude that the displacement driven by a power stroke is 8.3 nm. A working model of dimeric dynein driven by the 8-nm power stroke was proposed.
关键词： power stroke,optical tweezers,ATP,microtubule,dynein
更新于2025-09-23 15:21:21
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A novel intracellular signal amplification strategy for the quantification of ATP in single cells by microchip electrophoresis with laser induced fluorescence detection
摘要： An intracellular signal amplification strategy was developed for quantification of ATP in single cells by microchip electrophoresis with laser induced fluorescence detection. By using the method proposed, intracellular ATP levels in single Hela, HepG2 and HL-7702 cell were found in the range of 30～150, 30~140, and 19～120 fmol/cell, respectively.
关键词： laser induced fluorescence,ATP,microchip electrophoresis,single-cell analysis,signal amplification
更新于2025-09-23 15:21:01
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Use of BODIPY-Labeled ATP Analogues in the Development and Validation of a Fluorescence Polarization-Based Assay for Screening of Kinase Inhibitors
摘要： The screening of compound libraries to identify small-molecule modulators of specific biological targets is crucial in the process for the discovery of novel therapeutics and molecular probes. Considering the need for simple single-tool assay technologies with which one could monitor “all” kinases, we developed a fluorescence polarization (FP)-based assay to monitor the binding capabilities of protein kinases to ATP. We used BODIPY ATP-y-S as a probe to measure the shift in the polarization of a light beam when passed through the sample. We were able to optimize the assay using commercial Protein Kinase A (PKA) and H7 efficiently inhibited the binding of the probe when added to the reaction. Furthermore, we were able to employ the assay in a high-throughput fashion and validate the screening of a set of small molecules predicted to dock into the ATP-binding site of PKA. This will be useful to screen larger libraries of compounds that may target protein kinases by blocking ATP binding.
关键词： high-throughput screening,ATP-binding site,protein kinases,fluorescence polarization,BODIPY ATP-y-S
更新于2025-09-23 15:19:57
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Metabolic mapping with plasmonic nanoparticle assemblies
摘要： A rapid and simple methodology for the biomolecular analysis of single cells and microenvironments via a stick-and-peel plasmonic sensing platform is reported. Substrate-bound assemblies of plasmonic gold nanoparticles linked by reconfigurable oligonucleotides undergo disassembly upon target binding. Changes in the light scattering intensity of thousands of discrete nanoparticle assemblies are extrapolated concomitantly to yield the mapping of local target concentrations. The methodology is completely free of labelling, purification and separation steps. We quantified the intracellular ATP levels for two ovarian cancer cell lines to elucidate the differences and cellular distribution, and demonstrated the potential of the stick-and-peel platform for mapping the microenvironment of a 2D heterogeneous surface. The portable and economical analytical platform may broaden the affordability and applicability of single-cell based analyses and enable new opportunities in clinical care such as on-site molecular pathology.
关键词： stick-and-peel platform,plasmonic nanoparticle assemblies,ATP detection,single-cell analysis,molecular pathology
更新于2025-09-23 15:19:57
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Myosin II Filament Dynamics in Actin Networks Revealed with Interferometric Scattering Microscopy
摘要： The plasma membrane and the underlying cytoskeletal cortex constitute active platforms for a variety of cellular processes. Recent work has shown that the remodeling acto-myosin network modifies local membrane organization, but the molecular details are only partly understood because of difficulties with experimentally accessing the relevant time and length scales. Here, we use interferometric scattering microscopy to investigate a minimal acto-myosin network linked to a supported lipid bilayer membrane. Using the magnitude of the interferometric contrast, which is proportional to molecular mass, and fast acquisition rates, we detect and image individual membrane-attached actin filaments diffusing within the acto-myosin network and follow individual myosin II filament dynamics. We quantify myosin II filament dwell times and processivity as functions of ATP concentration, providing experimental evidence for the predicted ensemble behavior of myosin head domains. Our results show how decreasing ATP concentrations lead to both increasing dwell times of individual myosin II filaments and a global change from a remodeling to a contractile state of the acto-myosin network.
关键词： actin networks,ATP concentration,interferometric scattering microscopy,membrane organization,myosin II filament dynamics
更新于2025-09-23 15:19:57