21 results about "DAPI" patented technology

The invention discloses an image processing method and system based on an in-situ hybridization technology and a medium, and relates to the technical field of biological information, and the method comprises the following steps: converting a cell DAPI dyeing result into two-dimensional data from three-dimensional data by utilizing Z-axis maximum intensity projection, identifying and separating a single cell from the two-dimensional data by utilizing cell segmentation, and when the cell segmentation is used for processing an overlapping region, identifying and separating the single cell from the two-dimensional data. Using a registration algorithm Ashlar to calculate an error between adjacent visual fields of the same round to obtain cell position information; an imaging result after hybridization of the fluorescent probe and the gene is subjected to multiple rounds of registration through a registration algorithm Ashlar to generate a spliced image, local maximum values of the spliced image under different rounds are marked by using a fluorescent dot recognition algorithm, so that position information of each fluorescent dot is obtained, the position information of each fluorescent dot corresponds to the gene in a transcript, and the position information of each fluorescent dot corresponds to the gene in the transcript. Obtaining gene position information; distributing genes into cells by utilizing the cell position information and the gene position information to obtain a cell gene matrix and displaying the cell gene matrix; according to the image processing method and system and the medium, accurate matching of sequencing data and spatial information is achieved, and then the distribution rule of gene expression in space is better revealed.

The invention provides an organoid membrane protein and intracellular protein immunofluorescence in-situ co-staining method, which comprises the following steps of: sealing and fixing organoid glue drops, adding a mixed solution of membrane protein primary antibody and a Dune phosphate buffer solution containing bovine serum albumin for incubation, performing membrane permeation by using a permeation agent, sealing and washing, and drying to obtain the organoid membrane protein and intracellular protein immunofluorescence in-situ co-staining product. Adding a mixed solution of an intracellular protein primary antibody and a Dune phosphate buffer solution containing bovine serum albumin into the cell plate where the transparent organ-like glue drops are located, incubating and washing, then adding a mixed solution of a membrane protein and a fluorescent secondary antibody of the corresponding species of the intracellular protein, incubating and washing in a dark place, and finally obtaining the membrane protein / intracellular protein fluorescent antibody. And finally, adding a mixed solution of DAPI and a Duncheng phosphate buffer solution, and incubating and washing in a dark place again to obtain the membrane protein and intracellular protein immunofluorescence in-situ co-dyed organoid. According to the method, high-quality and high-fidelity membrane protein and intracellular protein in-situ co-dyeing is successfully realized, the integrity of a three-dimensional structure and the high efficiency of an experimental process are taken into consideration, and the method is convenient to operate, high in repeatability and good in universality.

The invention relates to the field of CAR-T cell concentration detection, in particular to a method for detecting CAR-T cells through combination of microfluidics and immunofluorescence. Through immunomagnetic bead enrichment and micro-fluidic chip partitioning, CAR-T cells are separated from a blood sample through immunomagnetic beads, then labeling is carried out or enhanced by adding a labeling antigen, anti-TCR-PE and DAPI, three-channel fluorescence labeling is achieved, quantitative analysis can be carried out on the number, the cell morphology and the function of the CAR-T cells through automatic immunofluorescence imaging, and the accuracy of CAR-T cell detection is improved. And the detection limit reaches 0.001%.

The invention discloses a method for predicting the curative effect of triple negative breast cancer immunotherapy by spatially quantizing CD39 + CD103 + CD8 + T cells, and relates to the technical field of biology.The method comprises the following specific steps that S1, an FFPE tumor tissue sample of a TNBC subject before immunotherapy is obtained, and a continuous section with the thickness of 3-5 microns is prepared and attached to a positive charge glass slide; s2, staining the section by using a combination containing CD8 / CD103 / CD39 / Pan-CK antibodies through a TSA sequential staining method, and then re-staining cell nucleuses by using DAPI; according to the method, spatial quantitative analysis is performed on a specific CD39 + CD103 + CD8 + T cell subset, a percentage index of target cells in CD8 + T cells and an average distance index of the target cells and nearest Pan-CK positive cells in a cancer nest region are set, and the target cells are determined according to comparison between the two indexes and a preset threshold value. According to the method, whether a subject is a potential benefit crowd treated by an anti-PD-1 / PD-L1 immune checkpoint inhibitor or not is accurately judged, a standardized technical means is provided for clinically screening the potential benefit crowd for immunotherapy, the accuracy and effectiveness of triple-negative breast cancer immunotherapy are effectively improved, and invalid treatment is reduced.

A spatial in situ sequencing method, belonging to the field of biology, is proposed. It utilizes an electric field-assisted directed migration of mRNA and in-situ capture with primers on a microarray surface to enrich tissue and release mRNA. In-situ reverse transcription generates covalently fixed cDNA, ensuring high positional stability during multiple rounds of hybridization and imaging. After reverse transcription, tissue is digested to remove tissue, reducing spatial hindrance and background interference, while the cDNA remains at its original coordinates due to covalent anchoring. Combining coding probe hybridization and RCA, single-molecule-level signal amplification and recognition are achieved. Through decoding and single-cell segmentation, transcripts are mapped to their respective cells, constructing a single-cell resolution spatial gene expression map. This method does not rely on multi-round DAPI mapping or other endogenous morphological marker-based multi-cycle image registration methods, making it suitable for high-throughput spatial in situ sequencing and significantly improving robustness and versatility in complex imaging scenarios such as thick tissue sections and low signal-to-noise ratios. It is applicable to high spatial resolution, high-throughput spatial transcriptome research.

The invention discloses a CCNE1 gene-based kit for evaluating the progress of bile duct cancer, belongs to the technical field of biomedical detection, and is used for evaluating the progress of bile duct cancer. The kit comprises a specific fluorescent probe, a hybridization buffer solution, a washing buffer solution, a positive reference substance, a DAPI dye solution and an anti-fluorescence quenching agent. Wherein the specific fluorescent probe is formed by coupling a fluorescent probe and amino-modified CCNE1 specific nucleic acid, and the fluorescent probe constructs a conjugated structure based on 5-methoxy-2, 3, 3-trimethylindole and 4-(9H-carbazole-9-yl) benzaldehyde. The CCNE1 gene expression level can be accurately reflected by detecting the intensity of fluorescence signals excited by 650 nm and emitted by 690 nm, then the malignant degree, metastasis potential and patient prognosis of the bile duct cancer are evaluated, the CCNE1 gene can be used for predicting the progression-free lifetime of a patient and the tumor metastasis risk, and a reliable tool is provided for diagnosis and treatment of the bile duct cancer.

The application provides a preparation kit for a Puccinellia distans cell nucleus suspension and relates to the field of molecular biology, and comprises a lysis buffer system, a phase separation system, a density gradient purification system and a lysis and staining system; wherein the lysis buffer system comprises 1xNIB working solution and 1xTAPS resuspension buffer, the pH value of the 1xTAPS resuspension buffer is 7.8-8.2; the phase separation system adopts ATPS two-phase separation solution; the density gradient purification system adopts 33%-37% Percoll separation solution; and the lysis and staining system comprises a cell lysis solution and a DAPI staining solution. The kit can obtain stable high-quality plant cell nucleus suspension, makes the cell nucleus damage small, improves the nucleus integrity rate, reduces the cell nucleus fragments and reduces the cell nucleus adhesion, and provides key technical support for plant single cell omics research of Puccinellia distans and the like.

The present application relates to the kiwi cultivation technical field, specifically to a method capable of early rapid detection of kiwi pollen activity, steps are as follows: S1, pollen sample is obtained at the early stage of kiwi male flower reproductive development; S2, anthers in pollen are soaked with Carnoy fixing solution; S3, DAPI staining solution is added to the soaked anthers, and a slice is prepared; S4, fluorescence microscope observation and activity determination are carried out.The present application has the beneficial effects that the present application can collect pollen and detect activity at the early stage of kiwi male flower reproductive development, and the activity detection result is consistent with that of the traditional method; subsequent accurate sampling is realized, work efficiency is improved, and comprehensive cost is reduced; pollination stability is ensured; pollen purity and collection efficiency are considered.

This invention discloses a method for detecting Candida albicans based on compound fluorescent staining and deep learning, belonging to the field of microbial detection technology. The method first synthesizes a novel NHC-triazolium salt fluorescent molecule, which is then compounded with calcium fluorescent white and DAPI to form a specific staining solution. After treating the sample with this staining solution, multi-channel fluorescence imaging is performed and images are acquired. Simultaneously, ITS sequencing is performed on the samples. Based on the sequencing results, the images are classified and bacterial cells are manually cropped to construct a labeled dataset. A convolutional neural network model is trained using this dataset to learn to recognize the unique "multi-fluorescent fingerprint" presented by Candida albicans stained with the compound staining solution. In application, the same staining and imaging are performed on the test samples, and the trained model automatically analyzes the images to achieve detection. This invention significantly improves the specificity of detection through the synergy of chemical staining and artificial intelligence. Experiments show that compared with traditional single staining or unstained methods, this method improves the specificity of Candida albicans recognition by 37% and 52%, respectively, and has the advantages of being fast, accurate, and highly automated.

The application belongs to the field of cell culture. The application provides a culture medium of rat amniotic membrane mesenchymal stem cells and a culture method of rat amniotic membrane mesenchymal stem cells. The culture medium is a complete culture medium, and the culture method is as follows: the rat amnion is cut into tissue blocks smaller than 2mm 2 , is uniformly laid in a culture dish with a spacing of 0.3-0.8cm, complete culture medium is slowly added after the tissue blocks are adhered, the culture medium is supplemented after 12h-36h, the primary medium replacement is performed after 48h-72h of culture, the impurity cells are removed, and the cells are subcultured when the cell fusion degree reaches 80%-90%. The light microscope observation and identification result show that the rat amniotic membrane mesenchymal stem cells cultured by the application basically conform to the cell morphology of rat amniotic membrane mesenchymal stem cells, the CD90&DAPI immunofluorescence identification result shows that the cell purity is greater than or equal to 90%, the cells have the characteristics of stem cells, and the culture method is efficient in amplification.

The invention discloses a DNA (Deoxyribose Nucleic Acid) probe ATP-3 and application thereof. The DNA (Deoxyribose Nucleic Acid) probe ATP-3 disclosed by the invention comprises DOX-3 and DOX-3-CP (Chlorinated Drug detection can be realized by using the DNA probe ATP-3 based on a non-label method DNA biosensor. Based on a double-strand competitive binding mechanism (DOX-3 and DOX specific binding leads to double-strand dissociation) of a DNA probe ATP-3, and in combination with a fluorescence switching characteristic (fluorescence enhancement during double-strand binding and quenching during dissociation) of an embedded fluorescent agent DAPI, trace detection of drugs is realized. By using a fluorospectro photometer as a detection auxiliary instrument, the method has the advantages of sensitive result, simplicity in operation, lower cost and the like.

The invention relates to a trace hydroxyputrescine lysine protein detection method, which comprises: 1) preparing a sample to be detected, and slicing cells, body fluid or tissue; (2) sealing the sample by using Duolink sealing liquid; (3) diluting an anti-rabbit PLUS probe and an anti-mouse MINUS probe Merck and DUO92101 / DUO92102 in proportion by using a Duolinkantibody diluent, incubating with a product obtained in the previous step, and respectively combining a rabbit polyclonal antibody and a mouse monoclonal antibody through the PLUS / MINUS probe to provide a basis for a ligation reaction; (4) diluting ligase by using a Duolink ligation buffer solution according to the ratio, and incubating with the product in the step (3) at the temperature; 5) diluting an amplification enzyme with a Duolink amplification buffer solution in proportion, and incubating with a product obtained in the previous step; 6, a mounting medium containing DAPI is adopted for mounting, and imaging analysis is carried out through a fluorescence or confocal microscopy. The problem that due to the fact that the affinity of the antibody is insufficient and the protein expression quantity is low, the omission ratio of the trace hydroxyputrescine lysine modified protein is high can be well solved.

The invention discloses a method for observing the shape and number of synapses in a dentate gyrus region based on image analysis, and relates to the technical field of neurobiology. The method comprises the following steps: S1, tissue treatment: after an experimental animal is anesthetized, pouring a buffer solution and a stationary solution into the heart in stages, separating hippocampus tissue, trimming and fixing, pre-cooling, slicing and preserving at low temperature; s2, dyeing and marking: performing targeted marking on the dentate gyrus area of the sea horse by adopting DiI particles through a fine tungsten steel through needle, and preserving at low temperature after incubation; s3, fixing and mounting: fixing for the second time, and mounting with an anti-fluorescence quenching agent after DAPI nuclear dyeing; s4, confocal imaging is carried out, exclusive parameter layer scanning is set, and qualified dendritic spine images are screened; and image quantitative analysis can be further carried out, and accurate quantification is realized according to classification standards. The method is standard in operation, greatly improves sample quality and experimental result credibility, optimizes experimental efficiency and controllability and reduces technical threshold by means of targeted marking and standardized processes, and is suitable for relevant basis and application research of the hippocampal synapses.

The invention provides a method for extracting, purifying and identifying fibroblasts, which comprises the following steps of: cutting meninx or spinal membrane tissues into pieces, digesting, and centrifuging to obtain a precipitate; re-suspending the obtained precipitate by using a fibroblast culture solution, spreading the precipitate in a polylysine pre-coated culture flask, culturing until the cell confluence degree is 90%, shaking, and discarding the supernatant liquid to obtain purified fibroblasts; and carrying out passage on the obtained purified fibroblasts, carrying out immobilization and permeabilization, sealing with goat serum, sequentially adding a primary antibody solution and a fluorescent secondary antibody solution, dyeing with DAPI, and observing and identifying the fibroblasts under a fluorescence microscope to complete the extraction, purification and identification of the fibroblasts. The invention also provides an application of the method. The method is used for extracting, purifying and identifying meningeal fibroblasts or spinal fibroblasts. The fibroblasts extracted by the method are high in purity, short in culture period and simple to operate.

The invention relates to the technical field of biomedicine, in particular to an automatic analysis and risk assessment system and method for peripheral blood endothelial progenitor cells, and the method comprises the following steps: collecting a peripheral blood sample and clinical data of a subject; the method comprises the following steps: enriching CD34 + cells from a peripheral blood sample based on microfluidic-immunomagnetic bead enrichment, carrying out VEGFR2 and DAPI immunofluorescent staining on the enriched cells, and obtaining a multi-channel fluorescent digital image; analyzing the multi-channel fluorescent digital image, identifying and counting VEGFR < 2 + > and DAPI < + > double-positive cells to obtain an EPC counting result; inputting the EPC counting result and clinical data of the subject into a multi-modal AI risk assessment model to obtain a risk assessment result; the health management suggestion information is formulated based on the risk assessment result, so that the model can have better suitability with the target subject, and the accuracy of risk assessment is improved.

The present application relates to pathological image processing technical field, provide a kind of tissue section image and DAPI staining image registration method, comprising: tissue section image and DAPI staining image are carried out gray processing, and according to same work magnification is enlarged;Obtain the feature point image of tissue section and DAPI staining;Find the best scaling ratio is 最佳 Of feature point image of tissue section and DAPI staining, rotation angle ia, horizontal displacement dx and vertical displacement dy and average point (xm, ym);With average point (xm, ym) generation multiple sub-frames, so that the tissue section feature point contained in sub-frame exceeds 200;For each sub-frame, using the best scaling ratio is 最佳 Rotation angle ia, in the case of (xm, ym) as rotation center, DAPI staining feature point is handled to generate transformation matrix tf2;Transformation matrix tf2 and DAPI staining image are generated final registration image by cv2.warpAffine function in opencv.Can tissue section image under normal microscope white light irradiation and DAPI staining image under fluorescence microscope are registered.

The invention discloses a flow ploidy analysis kit, and relates to the technical field of biological detection. According to the flow ploidy analysis kit provided by the invention, the DAPI staining solution is specifically combined with the DNA double-helix minor groove, so that accurate marking of the genetic material of the cell nucleus is realized, and the flow ploidy analysis kit can be directly used for flow ploidy analysis and detection of animal cells. By optimizing the working concentration of the DAPI staining solution, the toxic effect of the dye on cells is reduced to the maximum extent while the signal intensity is guaranteed. The kit is suitable for trace living samples, ploidy analysis can be completed without killing animals, and lossless monitoring support is provided for polyploidy breeding. The kit provided by the invention adopts DAPI dyeing, does not need an illumination activation step, can complete the dyeing process within 5 minutes of incubation at room temperature, has the advantage that a fluorescence signal is slightly influenced by pH fluctuation, has the advantage of'detection while dyeing ', and is especially suitable for rapid screening in a field or resource limited scene.

The invention provides a preparation kit of a pennisetum americanum cell nucleus suspension, and relates to the field of molecular biology, the preparation kit comprises a lysis buffer system, a phase separation system, a density gradient purification system and a lysis and dyeing system; wherein the lysis buffer solution system comprises a 1 * NIB working solution and a 1 * TAPS resuspension buffer solution, and the pH value of the 1 * TAPS resuspension buffer solution is 7.8-8.2; the phase separation system adopts ATPS two-phase separation liquid; the density gradient purification system adopts a Percoll separation solution with the concentration of 33%-37%; and the lysis and staining system comprises a cell lysis solution and a DAPI staining solution. The kit can obtain a stable high-quality plant cell nucleus suspension, so that cell nucleus damage is small, the nucleus integrity rate is improved, cell nucleus fragments are reduced, cell nucleus adhesion is reduced, and a key technical support is provided for single-cell omics research of plants such as pennisetum americana and the like.

The present invention discloses a method for extracting corn endosperm kernels suitable for ATAC-seq sequencing. The method comprises the following steps: (1) removing the seed coat and embryo from corn seeds, collecting the endosperm, and storing it at -80°C for a long time; taking approximately 5 g of the endosperm and manually grinding it into powder in liquid nitrogen; (2) adding 10 mL of sucrose buffer to each powder sample, mixing it thoroughly, and then filtering it through a 100 μm pore sieve; (3) passing the filtrate from step (2) through a 38 μm pore mesh, and then centrifuging the product at 700 × g for 5 minutes to collect the supernatant; (4) filtering the supernatant through a 38 μm sieve and centrifuging it at 2000 × g for 5 minutes. The cells were centrifuged at 1000 × g for 5 minutes, the supernatant was removed, and the precipitate was resuspended in 10 mL of sucrose buffer; (5) the cells were filtered through a 38 μm sieve again, centrifuged at 2000 × g for 5-10 minutes, the supernatant was removed, and the precipitate was resuspended in 5 mL of resuspension buffer; (6) the cells were centrifuged at 1000 × g for 5 minutes, the supernatant was discarded, the precipitate was suspended in 5 mL of PBS solution containing protease inhibitors, 10-20 μL of the solution was stained with DAPI at equal volumes for 10 minutes, and the cells were observed under a fluorescence microscope. The endosperm nuclei prepared by the present invention are numerous and highly active, which can overcome the influence of high starch content in storage tissues, provide strong technical support for subsequent single-cell sequencing, CUT&TAG, ATAC-seq and other related studies, and the extraction effect will also have an important impact on the experimental data.

This invention discloses a method and application for detecting goblet cell-related antigen channels in the piglet intestine. It relates to the field of antigen detection technology. The method includes the following steps: taking a portion of the mid-jejunum from a piglet; processing it; cutting the processed mid-jejunum into small pieces; incubating the pieces in a culture medium containing a marker; fixing; dehydrating and clearing the fixed pieces; embedding them in paraffin; preparing tissue sections; staining the tissue sections with a rabbit antibody against the goblet cell marker protein mucin 2 and a goat anti-rabbit fluorescent secondary antibody; mounting with the nuclear dye DAPI; and observing and analyzing under a fluorescence microscope. The method provided by this invention is simple to operate and can successfully detect goblet cell-related antigen channels in weaned piglets, laying the foundation for subsequent pig nutrition formulations.

This invention discloses a kit for assessing the progression of cholangiocarcinoma based on the CCNE1 gene, belonging to the field of biomedical detection technology, and is used to assess the progression of cholangiocarcinoma. The kit includes a specific fluorescent probe, hybridization buffer, washing buffer, positive control, DAPI staining solution, and an anti-fluorescence quencher. The specific fluorescent probe is formed by coupling a fluorescent probe with an amino-modified CCNE1-specific nucleic acid. The fluorescent probe is based on a conjugated structure constructed from 5-methoxy-2,3,3-trimethylindole and 4-(9H-carbazole-9-yl)benzaldehyde. By detecting the fluorescence signal intensity at 650 nm excitation and 690 nm emission, the expression level of the CCNE1 gene can be accurately reflected, thereby assessing the malignancy, metastatic potential, and patient prognosis of cholangiocarcinoma. It can also be used to predict progression-free survival and tumor metastasis risk, providing a reliable tool for the diagnosis and treatment of cholangiocarcinoma.