47 results about "Single molecule localization" patented technology

The invention relates to reconstructing a synthetic dense super-resolution image from at least one low-information-content image, for example from a sequence of diffraction-limited images acquired by single molecule localization microscopy. After having obtained such a sequence of diffraction-limited images, a sparse localization image is reconstructed from the obtained sequence of diffraction-limited images according to single molecule localization microscopy image processing. The reconstructed sparse localization image and/or a corresponding low-resolution wide-field image are input to an artificial neural network and a synthetic dense super-resolution image is obtained from the artificial neural network, the latter being trained with training data comprising triplets of sparse localization images, at least partially corresponding low-resolution wide-field images, and corresponding dense super-resolution images, as a function of a training objective function comparing dense super-resolution images and corresponding outputs of the artificial neural network.

A super-resolution optical imaging method based on a single-molecule positioning process is disclosed and used for observing an interaction process between a tumor cell exosome and a normal cell. The method includes (1) extracting the tumor cell exosome by utilizing a kit, (2) respectively labeling a membrane of the tumor cell exosome and a membrane surface receptor of the tumor cell exosome by using a first fluorescent molecule and a second fluorescent molecule, and staining the normal cell by using a third fluorescent molecule, and (3) observing the interaction process between the tumor cell exosome and the normal cell through a super-resolution optical microscope based on the single-molecule positioning process. The method overcomes the problem that exosomes cannot be observed carefully in the prior art due to a diffraction limit, and provides a novel technical means for researching exosome-mediated cancer metastasis mechanisms and for treating and researching cancer metastasis and spreading.

The invention discloses a real-time single-molecule positioning method guaranteeing precision and a system thereof. The method comprises the steps that region extraction is performed on an image to be processed based on pixel-level positioning parameters firstly; then coarse positioning is performed on the extracted image so that the subpixel-level positioning parameters of the extracted image are obtained; and then fine positioning is performed on the extracted image based on the subpixel-level positioning parameters of the extracted image. The initial value (the subpixel-level positioning parameters) obtained through coarse positioning is closer to a real value, based on which fine positioning calculation is performed so that the number of times of iteration in fine positioning is reduced, and thus positioning speed of a super-resolution image is accelerated. Therefore, positioning speed of the super-resolution image is accelerated under the condition of guaranteeing positioning precision.

The invention discloses a single-molecular positioning based fast super-resolution imaging method and system. The method comprises the following steps of: setting image acquisition parameters, hardware regulation and control parameters and super-resolution image processing and reconstruction parameters; performing hardware regulation and control according to the hardware regulation and control parameters; performing the super-resolution image processing and reconstruction on acquired images according to the super-resolution image processing and reconstruction parameters; and performing hardware regulation and control according to feedback results of super-resolution image processing and reconstruction. The single-molecular positioning based fast super-resolution imaging method adopts high algorithm precision, can be realized through parallel arithmetic of a graphic process unit (GPU), and can achieve higher image processing and reconstruction speed and automatically regulate and control the whole imaging system including a shutter, an electric bench and a step motor, thereby realizing fast super-resolution imaging.

The invention discloses a super-resolution optical imaging probe for living cells and a preparation method of the super-resolution optical imaging probe. Quantum dots are taken as a fluorophore, cell membrane penetrating peptides and nucleic acid aptamers are coupled on the surfaces of the quantum dots, the cell membrane penetrating peptides are used for promoting the probe to penetrate through the living cells, the nucleic acid aptamers are used for specific binding with nuclear membrane receptor protein of the living cells, and cell nucleus positioning of the probe is realized. The quantum dots with scintillating effects are taken as the fluorophore and are adapted to super-resolution optical imaging based on a single-molecule positioning method by means of the high luminous intensity and the fluorescence scintillation characteristic; meanwhile, the quantum dots with scintillating effects are taken as the fluorophore, a special imaging buffering solution is not required, therefore, the probe is adapted to super-resolution imaging of the living cells; besides, the preparation method is simple and convenient to operate, the cost is low, and the time cost and the economic cost are effectively saved.

A method of super-resolution microscopy is provided. The method includes mapping three-dimensional location of each single emission event in a plurality of single emission events from a series of optical images of a sample to create a point cloud. The point cloud is filtered or refined. Clusters within the point cloud are identified and characterized allowing for an assessment of molecular architecture.

The invention discloses a three-dimensional single-molecule positioning system based on a convolutional neural network. According to the system, a point spread function is calibrated by using a smallfluorescent ball sample; utilizing the point spread function images and a camera noise model to simulate and excite to generate training samples, and generating a true value three-dimensional matrix corresponding to each training image; inputting the training sample into an unsupervised noise reduction network to obtain noise reduction model parameters; inputting the denoised training sample and the true value three-dimensional matrix into a positioning neural network for training to obtain positioning model parameters; imaging the sample to be observed through a fluorescence microscope, and segmenting the image into the same size as the training set; performing noise reduction on the processed image; inputting the denoised image into the trained positioning network for testing; and finally, performing super-resolution reconstruction on an output result through a sparse coding method to obtain a super-resolution image. The system can maintain high precision and high accuracy of axial positioning for a point spread function excited by high overlapping rate and high density.

The invention discloses a super-resolution microscopic scale based on a quantum dot and DNA origami nano co-assembly structure, and belongs to the technical field of nano materials. The super-resolution microscopic scale is used for demarcating and calibrating the resolution of an ultrahigh-resolution fluorescence microscope system. According to the super-resolution microscopic scale based on thequantum dot and DNA origami nano co-assembly structure, quantum dots (2) and DNA single strands (3) are coupled and then modified to the fixed site of a DNA origami (1) (a triangle is taken as an example) to form the nano-scale with a fixed distance, so that the problem that the super-resolution microscopy lacks a high-precision scale to verify the accuracy of an obtained super-resolution image issolved. Through the super-resolution technology to break through the diffraction limit of light, the accuracy of the obtained super-resolution image is verified according to the fixed distance (20-200 nm) between the quantum dots on the same DNA origami structure. The super-resolution microscopic scale based on the quantum dot and DNA origami nano co-assembly structure is used for representing the imaging resolution of a positioning super-resolution monomolecular positioning microimaging technology (SMLM), and has the advantages of high brightness, stability, reusability and easiness in making different sizes.

The invention discloses an anti-bleaching unimolecule positioning three-dimensional super-resolution microscopy system. The anti-bleaching unimolecule positioning three-dimensional super-resolution microscopy system comprises a first laser, a beam expanding and collimating unit, a phase grating, a third lens, a first dichroic mirror, an objective lens, a sample table, an optical filter, a fourth lens, a detector, a second dichroic mirror and a second laser. The anti-bleaching unimolecule positioning three-dimensional super-resolution microscopy system utilizes a time domain focusing characteristic, fluorescence excitation of a sample is generated only on a time domain focusing plane, and meanwhile, fluorescence excitation in the whole time domain focusing plane can be achieved. Therefore,compared with the prior art, bleaching of a sample in a non-focusing plane can be avoided, and the working efficiency of SMS fluorescence microscopy technology can be effectively improved.

The invention discloses a manufacturing method and an application of an immunofluorescence co-localization imaging platform. A detection platform generates a gold nano array pattern through an electron beam lithography technology, and a surface is coupled with an HER2 (human epidermal growth factor receptor-2) nucleic acid aptamer with a fluorophore FAM through a gold sulfur bond. The platform canspecifically capture exosomes secreted by HER2 high-expression tumor cells; an imaging chain is an HER2 nucleic acid aptamer probe with another fluorophore Cy5; and two probes have fluorescence scintillation characteristics and can be used for super-resolution monomolecular positioning imaging (SMLM). Through an SMLM imaging technology, the captured exosomes can be subjected to super-resolution optical imaging; in combination with a double-color fluorescence co-localization technology, a false positive event can be judged at a single molecular level, and accuracy of a sandwich immunoassay technology is improved.

The invention discloses a telomere probe and a preparation method and application thereof. According to the telomere probe, a single stranded DNA is coupled onto the surface of the CdSSe/ZnS quantum dots, the telomere probe can be combined with the cell telomere by in-situ hybridization technique, and the telomere hybridized with the telomere probe is imaged by a single molecular positioning technique to realize the detection of the telomere. The telomere probe realizes the specific marking of the telomere, has higher resolution due to the adoption of the single molecule positioning microscopeimaging, and overcomes the limitation by the optical diffraction limit of the fluorescence microscopic technique. The telomere probe and the preparation method and the application thereof can be usedfor further study of the telomere structure.

The invention discloses a recombinant plasmid and cell line for monomolecular positioned super-resolution imaging of exosome and applications of the recombinant plasmid and the cell line. The recombinant plasmid is formed by fusing and constructing a fluorescence protein gene with a photoactivation property and a marking CD63 protein gene on an exosome film; Hela cells are transfected through a liposome-mediated plasmid transfection technology; and the Hela cells capable of stably expressing the photoactivation fluorescence protein marked exosome are obtained through drug screening. Monomolecular positioned super-resolution imaging researches of the separated and purified exosome and CD63 protein distribution in cells are carried out, with the help of the photoactivation fluorescence protein. Compared with a traditional microscope, the monomolecular positioned super-resolution imaging provides a better space resolution ratio and the monomolecular positioned super-resolution imaging ofthe exosome with activity is realized; and meanwhile, monomolecular positioned imaging researches of the CD63 protein distribution in the cells are also realized.

The invention discloses a single-molecule positioning micro-imaging method, an optical assembly and an imaging system. The method comprises the following steps: establishing an optical module with a double helix point dispersion function and a deformed multi-value pure phase grating multi-order imaging property; after fluorescence beams emitted by to-be-tested molecules of multiple sample surfacesare emitted through the optical module, carrying out imaging at different positions of the same detection surface, so as to respectively generate double helix images; determining transverse positionsof the to-be-tested molecules according to positions of the centers of double helix sidelobes in the double helix images on the imaging surfaces; and determining the axial positions of the to-be-tested molecules according to the middle points of the double helix sidelobes in the double helix images and a rotation angle of a connection line between the two sidelobes. The molecular information of multiple layers in the sample can be imaged at different positions of the same detection surface in a double helix manner, so that the axial positioning range and resolution ratio of a double helix point spread function can be improved without scanning, and the problem of large field depth detection in single molecule positioning and tracing techniques in living cells is solved.

Systems and methods assessing the accuracy of at least one localization from a single molecule localization microscopy (SMLM) dataset containing a plurality of localizations are described. The systems and methods calculate a Wasserstein-induced flux (WIF) value indicative of a confidence in the accuracy of a localization within the SMLM data set.

The invention discloses a super-resolution imaging probe based on silicon dioxide and a preparation method and application of the probe.The super-resolution imaging probe includes silicon dioxide nanoparticles connected with an Alexa Fluor 647 dye, and the surface of the super-resolution imaging probe is coupled with an HER2 (human epidermal growth factor receptor-2) nucleic acid aptamer; the probe can be used for targeted recognition of HER2 overexpression cell exosomes, that is, the HER2 overexpression cell exosomes are subjected to fluorescence labeling; super-resolution optical imaging canbe carried out on the cell exosome which specifically recognizes the probe by utilizing an ultra-high resolution microscope; the probe is uniform in particle size, small in size, high in positioningprecision, excellent in scintillation performance and suitable for ultrahigh-resolution optical imaging based on a single-molecule positioning method; and the exosome is marked by the super-resolutionimaging probe, and the exosome of the cell can be accurately marked by using a single-molecule positioning microscope in a double-channel co-positioning mode.

The invention discloses an intra-cellular miRNA quantifying method. The method comprises the following steps of (1) fixing excised cells, and then performing cell nucleus dyeing; (2) adding a buffer solution containing fluorescent probes to the cells, and enabling the fluorescent probes and intra-cellular miRNA to be detected to generate reversible combination; (3) placing the cells on an objective table of a TIRF microscope, performing detection, recording the shape of the cells at a bright field, performing fluorescence recording of the position of a cell nuclei at a wide field, and recording unimolecule fluorescence intensity-time track in an HILO mode; and (4) performing signal filtration to obtain a unimolecule imaging graph in which the unimolecule fluorescence intensity-time track presenting ON-OFF alternate changes, namely an intra-cellular miRNA unimolecule imaging graph, and performing counting so as to obtain the quantity of intra-cellular miRNA. According to the method disclosed by the invention, an "in-situ quantifying imaging" manner is provided, high-identification degree unimolecule dynamics fingerprint signals are used, unimolecule positioning and pinpoint countingare performed on the intra-cellular miRNA, and the technical bottleneck that intra-cellular miRNA quantifying depends on RNA extraction is broken through.

The invention discloses a feature determination method and device, electronic equipment and a storage medium, and belongs to the technical field of single molecule positioning imaging. The method comprises the following steps: constructing a Bezier curve according to feature information of sample points; determining a target sample image of the to-be-positioned image according to the point on the Bezier curve, the relationship between the sample point and the sample image and the to-be-positioned image; and taking the feature information of the sample points corresponding to the target sample image as the feature information of the points corresponding to the to-be-positioned image. According to the technical scheme, compared with an existing Gaussian fitting and cubic spline interpolation method, more imaging information is obtained, meanwhile, memory resources are reduced, and a new thought is provided for multi-dimensional single-molecule positioning imaging.