Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Super-resolution Geometric Barcoding for Multiplexed miRNA Profiling.

ABSTRACT: MicroRNA (miRNA) expression profiles hold promise as biomarkers for diagnostics and prognosis of complex diseases. Herein, we report a super-resolution fluorescence imaging-based digital profiling method for specific, sensitive, and multiplexed detection of miRNAs. In particular, we applied the DNA-PAINT (point accumulation for imaging in nanoscale topography) method to implement a super-resolution geometric barcoding scheme for multiplexed single-molecule miRNA capture and digital counting. Using synthetic DNA nanostructures as a programmable miRNA capture nano-array, we demonstrated high-specificity (single nucleotide mismatch discrimination), multiplexed (8-plex, 2 panels), and sensitive measurements on synthetic miRNA samples, as well as applied one 8-plex panel to measure endogenous miRNAs levels in total RNA extract from HeLa cells.

SUBMITTER: Xu W

PROVIDER: S-EPMC6314842 | biostudies-literature | 2018 Oct

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Super-resolution Geometric Barcoding for Multiplexed miRNA Profiling.

Xu Weidong W Yin Peng P Dai Mingjie M

Angewandte Chemie (International ed. in English) 20181004 43

MicroRNA (miRNA) expression profiles hold promise as biomarkers for diagnostics and prognosis of complex diseases. Herein, we report a super-resolution fluorescence imaging-based digital profiling method for specific, sensitive, and multiplexed detection of miRNAs. In particular, we applied the DNA-PAINT (point accumulation for imaging in nanoscale topography) method to implement a super-resolution geometric barcoding scheme for multiplexed single-molecule miRNA capture and digital counting. Usi ...[more]

PMID: 30182521

Similar Datasets

Demosaiced pixel super-resolution for multiplexed holographic color imaging.

Project description:To synthesize a holographic color image, one can sequentially take three holograms at different wavelengths, e.g., at red (R), green (G) and blue (B) parts of the spectrum, and digitally merge them. To speed up the imaging process by a factor of three, a Bayer color sensor-chip can also be used to demultiplex three wavelengths that simultaneously illuminate the sample and digitally retrieve individual set of holograms using the known transmission spectra of the Bayer color filters. However, because the pixels of different channels (R, G, B) on a Bayer color sensor are not at the same physical location, conventional demosaicing techniques generate color artifacts in holographic imaging using simultaneous multi-wavelength illumination. Here we demonstrate that pixel super-resolution can be merged into the color de-multiplexing process to significantly suppress the artifacts in wavelength-multiplexed holographic color imaging. This new approach, termed Demosaiced Pixel Super-Resolution (D-PSR), generates color images that are similar in performance to sequential illumination at three wavelengths, and therefore improves the speed of holographic color imaging by 3-fold. D-PSR method is broadly applicable to holographic microscopy applications, where high-resolution imaging and multi-wavelength illumination are desired.

| S-EPMC4926095 | biostudies-literature

Engineered fast-dissociating antibody fragments for multiplexed super-resolution microscopy.

Project description:Image reconstruction by integrating exchangeable single-molecule localization (IRIS) achieves multiplexed super-resolution imaging by high-density labeling with fast exchangeable fluorescent probes. However, previous methods to develop probes for individual targets required a great amount of time and effort. Here, we introduce a method for generating recombinant IRIS probes with a new mutagenesis strategy that can be widely applied to existing antibody sequences. Several conserved tyrosine residues at the base of complementarity-determining regions were identified as candidate sites for site-directed mutagenesis. With a high probability, mutations at candidate sites accelerated the off rate of recombinant antibody-based probes without compromising specific binding. We were able to develop IRIS probes from five monoclonal antibodies and three single-domain antibodies. We demonstrate multiplexed localization of endogenous proteins in primary neurons that visualizes small synaptic connections with high binding density. It is now practically feasible to generate fast-dissociating fluorescent probes for multitarget super-resolution imaging.

| S-EPMC9606137 | biostudies-literature

Fluorescence lifetime DNA-PAINT for multiplexed super-resolution imaging of cells.

Project description:DNA point accumulation for imaging in nanoscale topography (DNA-PAINT) is a powerful super-resolution technique highly suitable for multi-target (multiplexing) bio-imaging. However, multiplexed imaging of cells is still challenging due to the dense and sticky environment inside a cell. Here, we combine fluorescence lifetime imaging microscopy (FLIM) with DNA-PAINT and use the lifetime information as a multiplexing parameter for targets identification. In contrast to Exchange-PAINT, fluorescence lifetime PAINT (FL-PAINT) can image multiple targets simultaneously and does not require any fluid exchange, thus leaving the sample undisturbed and making the use of flow chambers/microfluidic systems unnecessary. We demonstrate the potential of FL-PAINT by simultaneous imaging of up to three targets in a cell using both wide-field FLIM and 3D time-resolved confocal laser scanning microscopy (CLSM). FL-PAINT can be readily combined with other existing techniques of multiplexed imaging and is therefore a perfect candidate for high-throughput multi-target bio-imaging.

| S-EPMC8752799 | biostudies-literature

Multiplexed 3D cellular super-resolution imaging with DNA-PAINT and Exchange-PAINT.

Project description:Super-resolution fluorescence microscopy is a powerful tool for biological research, but obtaining multiplexed images for a large number of distinct target species remains challenging. Here we use the transient binding of short fluorescently labeled oligonucleotides (DNA-PAINT, a variation of point accumulation for imaging in nanoscale topography) for simple and easy-to-implement multiplexed super-resolution imaging that achieves sub-10-nm spatial resolution in vitro on synthetic DNA structures. We also report a multiplexing approach (Exchange-PAINT) that allows sequential imaging of multiple targets using only a single dye and a single laser source. We experimentally demonstrate ten-color super-resolution imaging in vitro on synthetic DNA structures as well as four-color two-dimensional (2D) imaging and three-color 3D imaging of proteins in fixed cells.

| S-EPMC4153392 | biostudies-literature

Cancer cell profiling by barcoding allows multiplexed protein analysis in fine-needle aspirates.

Project description:Immunohistochemistry-based clinical diagnoses require invasive core biopsies and use a limited number of protein stains to identify and classify cancers. We introduce a technology that allows analysis of hundreds of proteins from minimally invasive fine-needle aspirates (FNAs), which contain much smaller numbers of cells than core biopsies. The method capitalizes on DNA-barcoded antibody sensing, where barcodes can be photocleaved and digitally detected without any amplification steps. After extensive benchmarking in cell lines, this method showed high reproducibility and achieved single-cell sensitivity. We used this approach to profile ~90 proteins in cells from FNAs and subsequently map patient heterogeneity at the protein level. Additionally, we demonstrate how the method could be used as a clinical tool to identify pathway responses to molecularly targeted drugs and to predict drug response in patient samples. This technique combines specificity with ease of use to offer a new tool for understanding human cancers and designing future clinical trials.

| S-EPMC4063286 | biostudies-literature

Ultrafast data mining of molecular assemblies in multiplexed high-density super-resolution images.

Project description:Multicolor single-molecule localization super-resolution microscopy has enabled visualization of ultrafine spatial organizations of molecular assemblies within cells. Despite many efforts, current approaches for distinguishing and quantifying such organizations remain limited, especially when these are contained within densely distributed super-resolution data. In theory, higher-order correlation such as the Triple-Correlation function is capable of obtaining the spatial configuration of individual molecular assemblies masked within seemingly discorded dense distributions. However, due to their enormous computational cost such analyses are impractical, even for high-end computers. Here, we developed a fast algorithm for Triple-Correlation analyses of high-content multiplexed super-resolution data. This algorithm computes the probability density of all geometric configurations formed by every triple-wise single-molecule localization from three different channels, circumventing impractical 4D Fourier Transforms of the entire megapixel image. This algorithm achieves 102-folds enhancement in computational speed, allowing for high-throughput Triple-Correlation analyses and robust quantification of molecular complexes in multiplexed super-resolution microscopy.

| S-EPMC6328550 | biostudies-literature

Automated highly multiplexed super-resolution imaging of protein nano-architecture in cells and tissues.

Project description:Understanding the nano-architecture of protein machines in diverse subcellular compartments remains a challenge despite rapid progress in super-resolution microscopy. While single-molecule localization microscopy techniques allow the visualization and identification of cellular structures with near-molecular resolution, multiplex-labeling of tens of target proteins within the same sample has not yet been achieved routinely. However, single sample multiplexing is essential to detect patterns that threaten to get lost in multi-sample averaging. Here, we report maS3TORM (multiplexed automated serial staining stochastic optical reconstruction microscopy), a microscopy approach capable of fully automated 3D direct STORM (dSTORM) imaging and solution exchange employing a re-staining protocol to achieve highly multiplexed protein localization within individual biological samples. We demonstrate 3D super-resolution images of 15 targets in single cultured cells and 16 targets in individual neuronal tissue samples with <10 nm localization precision, allowing us to define distinct nano-architectural features of protein distribution within the presynaptic nerve terminal.

| S-EPMC7096454 | biostudies-literature

Super-resolution ribosome profiling reveals unannotated translation events in Arabidopsis.

Project description:Deep sequencing of ribosome footprints (ribosome profiling) maps and quantifies mRNA translation. Because ribosomes decode mRNA every 3 nt, the periodic property of ribosome footprints could be used to identify novel translated ORFs. However, due to the limited resolution of existing methods, the 3-nt periodicity is observed mostly in a global analysis, but not in individual transcripts. Here, we report a protocol applied to Arabidopsis that maps over 90% of the footprints to the main reading frame and thus offers super-resolution profiles for individual transcripts to precisely define translated regions. The resulting data not only support many annotated and predicted noncanonical translation events but also uncover small ORFs in annotated noncoding RNAs and pseudogenes. A substantial number of these unannotated ORFs are evolutionarily conserved, and some produce stable proteins. Thus, our study provides a valuable resource for plant genomics and an efficient optimization strategy for ribosome profiling in other organisms.

| S-EPMC5111709 | biostudies-literature

Multiplexed and scalable super-resolution imaging of three-dimensional protein localization in size-adjustable tissues.

Project description:The biology of multicellular organisms is coordinated across multiple size scales, from the subnanoscale of molecules to the macroscale, tissue-wide interconnectivity of cell populations. Here we introduce a method for super-resolution imaging of the multiscale organization of intact tissues. The method, called magnified analysis of the proteome (MAP), linearly expands entire organs fourfold while preserving their overall architecture and three-dimensional proteome organization. MAP is based on the observation that preventing crosslinking within and between endogenous proteins during hydrogel-tissue hybridization allows for natural expansion upon protein denaturation and dissociation. The expanded tissue preserves its protein content, its fine subcellular details, and its organ-scale intercellular connectivity. We use off-the-shelf antibodies for multiple rounds of immunolabeling and imaging of a tissue's magnified proteome, and our experiments demonstrate a success rate of 82% (100/122 antibodies tested). We show that specimen size can be reversibly modulated to image both inter-regional connections and fine synaptic architectures in the mouse brain.

| S-EPMC5070610 | biostudies-literature

Multi-functional barcoding enables high resolution study of clonal dynamics [multiplexed HG3 ClonMapper recalls]

Project description:The remarkable evolutionary capacity of cancer poses major challenges to current therapeutic efforts, and results from the vast clonal heterogeneity and ability of individual cancer cells to adapt to diverse selective pressures. More complete mechanistic understanding of the basis of therapeutic resistance has been limited by difficulties in coupling information regarding genetic and epigenetic alterations present within individual cells to their respective transcriptomic and functional outputs. To this end, we developed a novel high-complexity expressed barcode system, ClonMapper, that integrates DNA barcoding with single-cell RNA-sequencing and clonal isolation to characterize thousands of clones within a mixed cancer cell population. In applying this system to the chronic lymphocytic leukemia cell line HG3 in the setting of resistance to fludarabine-based chemotherapy, we discover pretreatment sub-populations with distinct expression profiles (i.e. upregulated Wnt, Notch and CXCR4 signaling) that not only confer distinct treatment survivorship trajectories, but also provide the basis for long-term clonal equilibrium between co-existing clones in the absence of treatment. Characterization of individual clones comprising these sub-populations revealed remarkable genetic heterogeneity and the persistence of unique transcriptomic signatures throughout treatment exposure. These data reveal the diverse clonal characteristics and therapeutic responses of a heterogeneous cancer cell population and highlight the unprecedented resolution that can be achieved using ClonMapper.

2021-04-18 | GSE171833 | GEO

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