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Facile ultrasonic synthesized NH2-carbon quantum dots for ultrasensitive Co2+ ion detection and cell imaging
摘要： The amine decorated carbon quantum dots (NH2-CQDs) were synthesized through ultrasonic method from graphite rods derived CQDs and ammonia hydroxide and utilized as the sensing probes for cobalt (II) ions and nucleic acids. The sensing technique was investigated to be the fluorescence quenching effect, which demonstrated linear relationship between cobalt (II) ions concentration and the emission intensity deviation ratio in the concentration range of 50 nM to 40 μM with the detection limit of 12 nM. In brief, this sensitive and selective detection method was confirmed to demonstrate high potential in cobalt (II) ions detection in real samples and nucleic acid sensing in biological cells.
关键词： Nucleic acid sensing,Carbon quantum dots (CQDs),Cobalt sensor
更新于2025-11-19 16:56:42
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Digital LAMP on a Commercial Membrane
摘要： In this work, we report digital loop-mediated isothermal amplification (LAMP) or reverse-transcription LAMP (RT-LAMP) on a commercial membrane, without the need for complex chip fabrication or use of specialized equipment. Due to the pore size distribution, the theoretical error for digital LAMP on these membranes was analyzed, using a combination of Random Distribution Model and Multi-volume Theory. A facile peel-off process was developed for effective droplets formation on the commercial track-etched polycarbonate (PCTE) membrane. Each pore functions as an individual nanoreactor for single DNA amplification. Absolute quantification of bacteria genomic DNA was realized with a dynamic range from 11 to 1.1 × 10^5 copies/μL. One-step digital RT-LAMP was also successfully performed on the membrane for the quantification of MS2 virus in wastewater. With the introduction of new probes, the positive pores can be easily distinguished from negative ones with 100 times difference in fluorescence intensities. Finally, the cost of a disposable membrane is less than $0.1/piece, which, to the best of our knowledge, is the most inexpensive way to perform digital LAMP. The membrane system offers opportunities for point-of-care users or common laboratories to perform digital quantification, single cell analysis, or other bioassays in an inexpensive, flexible and simplified way.
关键词： Digital LAMP,RT-LAMP,PCR,Droplets,Virus,Microfluidic,Membrane,Nucleic acid
更新于2025-09-23 15:23:52
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Peptide Nucleic Acid–Fluorescence In Situ Hybridization for Detection of Staphylococci From Endophthalmitis Isolates: A Proof-of-Concept Study
摘要： PURPOSE. Rapid identi?cation of pathogens causing endophthalmitis may improve treatment outcomes through early administration of species-speci?c medication. The current study reports a new molecular application of peptide nucleic acid–?uorescence in situ hybridization (PNA-FISH) with Staphylococcus-speci?c molecular PNA probes for the potential rapid detection of common pathogens causing endophthalmitis. METHODS. An experimental study was designed to evaluate the proof of concept at the microbiology laboratory of the Bascom Palmer Eye Institute. Stored culture-positive staphylococci endophthalmitis isolates obtained from prior vitreous samples (n ? 15), along with broth as negative controls (n ? 5) were used. Inoculum was prepared to a ?nal concentration of 1 3 105 colony-forming units/mL to ensure that the isolates were viable. Smears of samples were ?xed and hybridized using QuickFISH protocol with probes for Staphylococcus. RESULTS. With PNA-FISH technique, Staphylococcus aureus was identi?ed in 9 of 10 samples and coagulase-negative staphylococci were identi?ed in 10 of 10 samples. Detection time was 20 minutes. CONCLUSIONS. This study serves a proof of concept using a new microbial detection system with FISH probes, and may have the potential for clinical use in the rapid and accurate identi?cation of isolates from patients with endophthalmitis.
关键词： peptide nucleic acid (PNA),endophthalmitis,rapid identification,fluorescence in situ hybridization (FISH)
更新于2025-09-23 15:22:29
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Integration of plasmonic heating and ona??chip temperature sensor for nucleic acid amplification assays
摘要： Nucleic acid tests have been widely used for diagnosis of diseases by detecting the relevant genetic markers that are usually amplified using polymerase chain reaction (PCR). This work reports the use of a plasmonic device as an efficient and low-cost PCR thermocycler to facilitate nucleic acid-based diagnosis. The thermoplasmonic device, consisting of a one-dimensional metal grating, exploited the strong light absorption of plasmonic resonance modes to heat up PCR reagents using a near-infrared laser source. The plasmonic device also integrated a thin-film thermocouple on the metal grating to monitor the sample temperature. The plasmonic thermocycler is capable of performing a PCR amplification cycle in approximately 2.5 minutes. We successfully demonstrated the multiplex and real-time PCR amplifications of the antibiotic resistance genes using the genomic DNAs extracted from Acinetobacter baumannii, Klebsiella pneumonia, Escherichia coli, and Campylobacter.
关键词： polymer chain reaction,thermoplasmonics,thermocycler,nucleic acid-based diagnostics,antibiotic resistance detection,Photothermal effect
更新于2025-09-23 15:19:57
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Rapid Quantitative Fluorescence Detection of Copper Ions with Disposable Microcapsule Arrays Utilizing Functional Nucleic Acid Strategy
摘要： In this work, an economical and easy-to-use microcapsule array fabricated by ice printing technique has been realized for ultrasensitive fluorescence quantification of copper ions employing functional nucleic acid strategy. With ice printing, the detection reagents are sealed by polystyrene (PS) film isolation and photopolymer, which guarantees a stable and contamination-free environment for functional nucleic acid reaction. Our microcapsule arrays have shown long-term stability (20 days) under ?20 °C storage in frozen form before use. During the Cu2+ on-site detection, 1 μL sample is simply injected into the thawy microcapsule by a microliter syringe under room temperature, and after 20 minutes the fluorescence result can be obtained by an LED transilluminator. This method can realize the detection limit to 100 nM (100 fmol/μL) with high specificity.
关键词： ice printing,functional nucleic acid,fluorescence detection,copper ions,microcapsule arrays
更新于2025-09-19 17:15:36
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Colorimetric Polymerase Chain Reaction Enabled by a Fast Light-Activated Substrate Chromogenic Detection Platform
摘要： Miniaturization of nucleic acid tests (NATs) into portable, inexpensive detection platforms may aid disease diagnosis in point-of-care (POC) settings. Colorimetric signals are ideal readouts for portable NATs, and it remains of high demand to develop color readouts that are simple, quantitative and versatile. Thus motivated, we report a Fast Light-Activated Substrate cHromogenic polymerase chain reaction (FLASH PCR) that uses DNA intercalating dyes (DIDs) to enable colorimetric nucleic acid detection and quantification. The FLASH system is established on our finding that DID-DNA intercalation can promote the rapid photooxidation of chromogenic substrates through light-induced production of singlet oxygen. Using this principle, we have successfully converted DID-based fluorescent PCR assays into colorimetric FLASH PCR. To demonstrate the practical applicability of FLASH PCR to POC diagnosis, we also fabricated two readout platforms, including a portable electronic FLASH reader and a paper-based FLASH strip. Using the FLASH reader, we were able to detect as low as 60 copies of DNA standards, a limit of detection (LOD) comparable with commercial quantitative PCR. The FLASH strip further enables the reader-free detection of PCR amplicons by converting the colorimetric signal into the visual measurement of distance as a readout. Finally, the practical applicability of the FLASH PCR was demonstrated by the detection and/or quantification of nucleic acid markers in diverse clinical and biological samples.
关键词： FLASH,Singlet Oxygen,Photooxidation,DNA Intercalating Dyes,Colorimetric,Nucleic Acid Tests,Paper-based Strip,Point-of-Care,Portable Electronic Reader,Polymerase Chain Reaction
更新于2025-09-19 17:13:59
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Immobilization of ssDNA on the surface of silver nanoparticles-graphene quantum dots modified by gold nanoparticles towards biosensing of microorganism
摘要： Rapid screening of pathogenic microorganisms are widely demand in the last decade. According to fastidious and hard to grow nature, Legionella penumophila is important pathogen for bioassay and biomedical analysis. In this work a novel paper-based genoassay was developed for monitoring of L. penumophila. For the first time, silver-graphene quantum dots (Ag/GQDs) ink was synthesized and used for construction of new substrate for bioassay of L. penumophila. The prepared interface was modified by gold nanoparticles grafted by Cysteamine A (CysA/AuNPs) which is necessary for ssDNA immobilization and hybridization by cDNA. All of the genoassay fabrication steps were characterized by field emission scanning electron microscope (FE-SEM), Energy-dispersive X-ray spectroscopy (EDS), Atomic force microscopic (AFM) and also TEM (transmission electron microscopy). Using chronoamperometry technique, the measurement of target cDNA was performed successfully. Also, cDNA was determined in the linear rang of 1μM to 1ZM which low limit of quantification was 1ZM. The results show that the designed bio-platform despite a simple structure with high sensitivity and specificity for the DNA based bioassay of L. penumophila genome.
关键词： nanotechnology,hybridization,biosensor,nucleic acid,advanced nanomaterial,bioanalysis
更新于2025-09-11 14:15:04
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Quantitative fluorescence imaging determines the absolute number of locked nucleic acid oligonucleotides needed for suppression of target gene expression
摘要： Locked nucleic acid based antisense oligonucleotides (LNA-ASOs) can reach their intracellular RNA targets without delivery modules. Functional cellular uptake involves vesicular accumulation followed by translocation to the cytosol and nucleus. However, it is yet unknown how many LNA-ASO molecules need to be delivered to achieve target knock down. Here we show by quantitative fluorescence imaging combined with LNA-ASO microinjection into the cytosol or unassisted uptake that ～105 molecules produce >50% knock down of their targets, indicating that a substantial amount of LNA-ASO escapes from endosomes. Microinjected LNA-ASOs redistributed within minutes from the cytosol to the nucleus and remained bound to nuclear components. Together with the fact that RNA levels for a given target are several orders of magnitude lower than the amounts of LNA-ASO, our data indicate that only a minor fraction is available for RNase H1 mediated reduction of target RNA. When non-specific binding sites were blocked by co-administration of non-related LNA-ASOs, the amount of target LNA-ASO required was reduced by an order of magnitude. Therefore, dynamic processes within the nucleus appear to influence the distribution and activity of LNA-ASOs and may represent important parameters for improving their efficacy and potency.
关键词： RNase H1,Locked nucleic acid,nuclear accumulation,microinjection,gene knock down,antisense oligonucleotides,quantitative fluorescence imaging
更新于2025-09-10 09:29:36
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A Rapid, Label-free and Impedimetric DNA Sensor Based on PNA-modified Nanoporous Gold Electrode
摘要： A gold nanoparticle film was successfully prepared on highly ordered and porous anodic aluminum oxide substrate through thermal evaporation deposition technique. As-fabricated nanoporous gold electrode modified by thiol-derivative peptide nucleic acid (PNA) was applied for impedimetric DNA sensing. In-situ kinetic analysis of PNA/DNA hybridization process was realized by electrochemical impedance spectroscopy (EIS) method. Considering the special structure of nanoporous gold electrode, a new equivalent circuit model was proposed for EIS data fitting. The sensitivity of PNA-modified nanoporous gold electrode is 0.4 Ω/cm2·nM. Meanwhile, complementary DNA target could be detected at concentration as low as 10 nM, which indicating that a methodology for rapid, sensitive and label-free detection of DNA has been set-up based on nanoporous gold electrode.
关键词： DNA sensing,nanoporous gold electrode,peptide nucleic acid,anodic aluminum oxide
更新于2025-09-10 09:29:36
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Visualized Quantitation of Trace Nucleic Acids Based on Coffee-Ring Effect on Colloid-Crystal Substrate
摘要： We report a visualized quantitative detection method for nucleic-acid amplification tests based on coffee-ring effect on colloid-crystal substrate. The solution for loop-mediated isothermal amplification (LAMP) of DNA is dropcast on a colloid-crystal surface. After complete drying, a coffee ring containing the LAMP byproduct (i.e. magnesium pyrophosphate) is formed, and it is found that the width of the coffee ring is linearly correlated to the logarithm of the original DNA concentration before the isothermal amplification. Importantly, compared with other substrates, we found that the colloidal-crystal substrate is an appropriate substrate for carrying out the assay of high sensitivity. Based on these findings, we develop a coffee ring-based assay for quantitative readout of trace DNA in sample. The assay requires 0.50 μL sample and is completed in 5 min in a homemade chamber with constant humidity. Semi-quantitative detection of trace DNA is performed using naked eyes. With the use of a smartphone, the DNA in a sample can be quantitatively detected with a limit-of-detection of 20 copies.
关键词： nucleic-acid amplification tests,colloid-crystal substrate,LAMP,quantitative detection,coffee-ring effect
更新于2025-09-10 09:29:36