Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Single-Molecule Detection of Nucleic Acids via Liposome Signal Amplification in Mass Spectrometry.

ABSTRACT: A single-molecule detection method was developed for nucleic acids based on mass spectrometry counting single liposome particles. Before the appearance of symptoms, a negligible amount of nucleic acids and biomarkers for the clinical diagnosis of the disease were already present. However, it is difficult to detect extremely low concentrations of nucleic acids using the current methods. Hence, the establishment of an ultra-sensitive nucleic acid detection technique is urgently needed. Herein, magnetic beads were used to capture target nucleic acids, and liposome particles were employed as mass tags for single-particle measurements. Liposomes were released from magnetic beads via photocatalytic cleavage. Hence, one DNA molecule corresponded to one liposome particle, which could be counted using mass spectrometric measurement. The ultrasensitive detection of DNA (10-18 M) was achieved using this method.

SUBMITTER: Lin X

PROVIDER: S-EPMC8963037 | biostudies-literature | 2022 Feb

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Single-Molecule Detection of Nucleic Acids via Liposome Signal Amplification in Mass Spectrometry.

Lin Xiangcheng X Zhao Mengmeng M Li Mingyue M Long Juan J Zhang Jing J Yu Fang F Xu Fen F Sun Lixian L

Sensors (Basel, Switzerland) 20220210 4

A single-molecule detection method was developed for nucleic acids based on mass spectrometry counting single liposome particles. Before the appearance of symptoms, a negligible amount of nucleic acids and biomarkers for the clinical diagnosis of the disease were already present. However, it is difficult to detect extremely low concentrations of nucleic acids using the current methods. Hence, the establishment of an ultra-sensitive nucleic acid detection technique is urgently needed. Herein, mag ...[more]

PMID: 35214249

Similar Datasets

Single molecule mass photometry of nucleic acids.

Project description:Mass photometry is a recently developed methodology capable of measuring the mass of individual proteins under solution conditions. Here, we show that this approach is equally applicable to nucleic acids, enabling their facile, rapid and accurate detection and quantification using sub-picomoles of sample. The ability to count individual molecules directly measures relative concentrations in complex mixtures without need for separation. Using a dsDNA ladder, we find a linear relationship between the number of bases per molecule and the associated imaging contrast for up to 1200 bp, enabling us to quantify dsDNA length with up to 2 bp accuracy. These results introduce mass photometry as an accurate, rapid and label-free single molecule method complementary to existing DNA characterization techniques.

| S-EPMC7515692 | biostudies-literature

A mutation-sensitive, multiplexed and amplification-free detection of nucleic acids by stretching single-molecule tandem hairpin probes.

Project description:The detection of nucleic acid sequences in parallel with the discrimination of single nucleotide variations (SNVs) is critical for research and clinical applications. A few limitations make the detection technically challenging, such as too small variation in probe-hybridization energy caused by SNVs, the non-specific amplification of false nucleic acid fragments and the few options of dyes limited by spectral overlaps. To circumvent these limitations, we developed a single-molecule nucleic acid detection assay without amplification or fluorescence termed THREF (hybridization-induced tandem DNA hairpin refolding failure) based on multiplexed magnetic tweezers. THREF can detect DNA and RNA sequences at femtomolar concentrations within 30 min, monitor multiple probes in parallel, quantify the expression level of miR-122 in patient tissues, discriminate SNVs including the hard-to-detect G-U or T-G wobble mutations and reuse the probes to save the cost. In our demonstrative detections using mock clinic samples, we profiled the let-7 family microRNAs in serum and genotyped SARS-CoV-2 strains in saliva. Overall, the THREF assay can discriminate SNVs with the advantages of high sensitivity, ultra-specificity, multiplexing, reusability, sample hands-free and robustness.

| S-EPMC10516651 | biostudies-literature

Peptide nucleic acids as tools for single-molecule sequence detection and manipulation.

Project description:The ability to strongly and sequence-specifically attach modifications such as fluorophores and haptens to individual double-stranded (ds) DNA molecules is critical to a variety of single-molecule experiments. We propose using modified peptide nucleic acids (PNAs) for this purpose and implement them in two model single-molecule experiments where individual DNA molecules are manipulated via microfluidic flow and optical tweezers, respectively. We demonstrate that PNAs are versatile and robust sequence-specific tethers.

| S-EPMC3322611 | biostudies-literature

Towards single-molecule nanomechanical mass spectrometry.

Project description:Mass spectrometry provides rapid and quantitative identification of protein species with relatively low sample consumption. The trend towards biological analysis at increasingly smaller scales, ultimately down to the volume of an individual cell, continues, and mass spectrometry with a sensitivity of a few to single molecules will be necessary. Nanoelectromechanical systems provide unparalleled mass sensitivity, which is now sufficient for the detection of individual molecular species in real time. Here, we report the first demonstration of mass spectrometry based on single biological molecule detection with a nanoelectromechanical system. In our nanoelectromechanical-mass spectrometry system, nanoparticles and protein species are introduced by electrospray injection from the fluid phase in ambient conditions into vacuum, and are subsequently delivered to the nanoelectromechanical system detector by hexapole ion optics. Precipitous frequency shifts, proportional to the mass, are recorded in real time as analytes adsorb, one by one, onto a phase-locked, ultrahigh-frequency nanoelectromechanical resonator. These first nanoelectromechanical system-mass spectrometry spectra, obtained with modest mass sensitivity from only several hundred mass adsorption events, presage the future capabilities of this approach. We also outline the substantial improvements that are feasible in the near term, some of which are unique to nanoelectromechanical system based-mass spectrometry.

| S-EPMC3846395 | biostudies-literature

Compaction of Duplex Nucleic Acids upon Native Electrospray Mass Spectrometry.

Project description:We report on the fate of nucleic acids conformation in the gas phase as sampled using native mass spectrometry coupled to ion mobility spectrometry. On the basis of several successful reports for proteins and their complexes, the technique has become popular in structural biology, and the conformation survival becomes more and more taken for granted. Surprisingly, we found that DNA and RNA duplexes, at the electrospray charge states naturally obtained from native solution conditions (?100 mM aqueous NH4OAc), are significantly more compact in the gas phase compared to the canonical solution structures. The compaction is observed for all duplex sizes (gas-phase structures are more compact than canonical B-helices by ?20% for 12-bp, and by up to ?30% for 36-bp duplexes), and for DNA and RNA alike. Molecular modeling (density functional calculations on small helices, semiempirical calculations on up to 12-bp, and molecular dynamics on up to 36-bp duplexes) demonstrates that the compaction is due to phosphate group self-solvation prevailing over Coulomb repulsion. Molecular dynamics simulations starting from solution structures do not reproduce the experimental compaction. To be experimentally relevant, molecular dynamics sampling should reflect the progressive structural rearrangements occurring during desolvation. For nucleic acid duplexes, the compaction observed for low charge states results from novel phosphate-phosphate hydrogen bonds formed across both grooves at the very late stages of electrospray.

| S-EPMC5445532 | biostudies-literature

Utilization of chromogenic enzyme substrates for signal amplification in multiplexed detection of biomolecules using surface mass spectrometry.

Project description:MicroRNAs (miRNAs) are important post-transcriptional gene regulators and can serve as potential biomarkers for many diseases. Most of the current miRNA detection techniques require purification from biological samples, amplification, labeling, or tagging, which makes quantitative analysis of clinically relevant samples challenging. Here we present a new strategy for the detection of miRNAs with uniformity over a large area based on signal amplification using enzymatic reactions and measurements using time-of-flight secondary ion mass spectrometry (ToF-SIMS), a sensitive surface analysis tool. This technique has high sequence specificity through hybridization with a hairpin DNA probe and allows the identification of single-base mismatches that are difficult to distinguish by conventional mass spectrometry. We successfully detected target miRNAs in biological samples without purification, amplification, or labeling of target molecules. In addition, by adopting a well-known chromogenic enzymatic reaction from the field of biotechnology, we extended the use of enzyme-amplified signal enhancement ToF (EASE-ToF) to protein detection. Our strategy has advantages with respect to scope, quantification, and throughput over the currently available methods, and is amenable to multiplexing based on the outstanding molecular specificity of mass spectrometry (MS). Therefore, our technique not only has the potential for use in clinical diagnosis, but also provides evidence that MS can serve as a useful readout for biosensing to perform multiplexed analysis extending beyond the limitations of existing technology.

| S-EPMC7845929 | biostudies-literature

Detection of nucleic acids <i>via</i> G-quadruplex-controlled l-cysteine oxidation and catalyzed hairpin assembly-assisted signal amplification.

Project description:The development of simple, sensitive and cost-effective methods for specific nucleic acid detection has attracted tremendous attention due to its importance to the early diagnosis of genetic diseases and to biodefense applications. In this work, we demonstrated a fluorescent turn-off mode DNA assay based on l-cysteine-modulated synthesis of CdTe quantum dots (CdTe QDs), horseradish peroxidase-mimicking G-quadruplex-hemin-K+ complex controlled oxidation of l-cysteine to cystine, and catalyzed hairpin assembly (CHA)-assisted signal amplification. After the addition of target DNA, the CHA signal amplification reaction was triggered and numerous H1-H2 double-stranded DNA were formed, initiating the release of G-quadruplex sequences in H2 simultaneously. Thus, the degree of inhibition of the synthesis of CdTe QDs is proportional to the concentration of the G-quadruplex sequence in this method. In contrast, when the target DNA was absent, the CHA could not be triggered, and the fluorescence signal was high due to the remaining intact l-cysteine. Under optimal experimental conditions, the homogeneous fluorescence method achieved the detection of HIV DNA with a linear range from 0.1 pM to 1 nM and a detection limit of 0.12 pM. This novel biosensor exhibits excellent specificity in differentiating DNA sequences with a single-base and two-base mismatch. To the best of our knowledge, this a label-free and highly sensitive bioassay utilizing CHA-assisted signal amplification and G-quadruplex control of in situ synthesis of CdTe QDs strategy was not reported in previous. Thus, this proposed strategy is anticipated to find use in basic biochemical research and clinical diagnosis.

| S-EPMC9091423 | biostudies-literature

Ultra-sensitive and rapid detection of nucleic acids and microorganisms in body fluids using single-molecule tethering.

Project description:Detection of microbial nucleic acids in body fluids has become the preferred method for rapid diagnosis of many infectious diseases. However, culture-based diagnostics that are time-consuming remain the gold standard approach in certain cases, such as sepsis. New culture-free methods are urgently needed. Here, we describe Single MOLecule Tethering or SMOLT, an amplification-free and purification-free molecular assay that can detect microorganisms in body fluids with high sensitivity without the need of culturing. The signal of SMOLT is generated by the displacement of micron-size beads tethered by DNA probes that are between 1 and 7 microns long. The molecular extension of thousands of DNA probes is determined with sub-micron precision using a robust and rapid optical approach. We demonstrate that SMOLT can detect nucleic acids directly in blood, urine and sputum at sub-femtomolar concentrations, and microorganisms in blood at 1 CFU mL-1 (colony forming unit per milliliter) threefold faster, with higher multiplexing capacity and with a more straight-forward protocol than amplified methodologies. SMOLT's clinical utility is further demonstrated by developing a multiplex assay for simultaneous detection of sepsis-causing Candida species directly in whole blood.

| S-EPMC7508858 | biostudies-literature

A Sensitive Potentiometric Sensor for Isothermal Amplification-Coupled Detection of Nucleic Acids.

Project description:A disposable potentiometric sensor was newly developed for the amplification-coupled detection of nucleic acids. The hydrogen-ion is generally released during isothermal amplification of nucleic acids. The surface potential on the oxide-functionalized electrode of the extended gate was directly measured using full electrical circuits with the commercial metal-oxide semiconductor field-effect transistors (MOSFETs) and ring oscillator components, which resulted in cost-effective, portable and scalable real-time nucleic acid analysis. The current-starved ring oscillator changes surface potential to its frequency depending on the square of the variation in pH with a high signal-to-noise ratio during isothermal amplification. The device achieves a conversion rate of 20.5 kHz/mV and a detection resolution of 200 µV for the surface potential. It is demonstrated that the sensor successfully monitors in real-time isothermal amplification of the extracted nucleic acids from Salmonella pathogenic bacteria. The in situ variations in the frequency of the pH-sensitive sensor were compared with the results of both a conventional optical device and pH-meter during isothermal amplification.

| S-EPMC6068556 | biostudies-literature

Tyramide signal amplification mass spectrometry (TSA-MS) ratio identifies nuclear speckle proteins.

Project description:We present a simple ratio method to infer protein composition within cellular structures using proximity labeling approaches but compensating for the diffusion of free radicals. We used tyramide signal amplification (TSA) and label-free mass spectrometry (MS) to compare proteins in nuclear speckles versus centromeres. Our "TSA-MS ratio" approach successfully identified known nuclear speckle proteins. For example, 96% and 67% of proteins in the top 30 and 100 sorted proteins, respectively, are known nuclear speckle proteins, including proteins that we validated here as enriched in nuclear speckles. We show that MFAP1, among the top 20 in our list, forms droplets under certain circumstances and that MFAP1 expression levels modulate the size, stability, and dynamics of nuclear speckles. Localization of MFAP1 and its binding partner, PRPF38A, in droplet-like nuclear bodies precedes formation of nuclear speckles during telophase. Our results update older proteomic studies of nuclear speckles and should provide a useful reference dataset to guide future experimental dissection of nuclear speckle structure and function.

| S-EPMC7480118 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data