24 results about "Blot" patented technology

The application belongs to the technical field of biology and particularly relates to an African swine fever virus pNP868R protein monoclonal antibody and a preparation method and application thereof. The application takes the pNP868R protein prepared by eukaryotic expression as an antigen, immunizes BALB / c mice, uses hybridoma technology, and obtains a hybridoma cell line capable of stably secreting the anti-African swine fever virus pNP868R protein monoclonal antibody through cell fusion, indirect ELISA method screening, limited dilution method subcloning and hybridoma continuous passage culture. The application also obtains an African swine fever virus pNP868R protein monoclonal antibody. The monoclonal antibody has high affinity with the pNP868R protein and can be applied in Western-Blot experiments, indirect immunofluorescence experiments, immunoprecipitation experiments and the capture of proteins interacting with pNP868R, and can be used for the diagnosis, prevention or treatment of African swine fever.

This invention discloses an anti-MSLN antibody and its applications. The anti-MSLN antibody comprises a heavy chain variable region and a light chain variable region. The heavy chain variable region comprises HCDR1, HCDR2, and HCDR3, and the light chain variable region comprises LCDR1, LCDR2, and LCDR3. The amino acid sequences of HCDR1, HCDR2, and HCDR3 are shown in SEQ ID NO:7, 8, and 9, respectively; and the amino acid sequences of LCDR1, LCDR2, and LCDR3 are shown in SEQ ID NO:4, 5, and 6, respectively. The antibody provided by this invention has good affinity and specificity for MSLN antigens, effectively detecting target proteins in cells overexpressing MSLN. It can also be widely used as a core ingredient in the development of high-sensitivity MSLN detection kits such as ELISA, Western blotting, and immunohistochemistry.

This invention discloses an anti-MSLN antibody and its applications. The anti-MSLN antibody comprises a heavy chain variable region and a light chain variable region. The heavy chain variable region comprises HCDR1, HCDR2, and HCDR3, and the light chain variable region comprises LCDR1, LCDR2, and LCDR3. The amino acid sequences of HCDR1, HCDR2, and HCDR3 are shown in SEQ ID NO:7, 8, and 9, respectively; and the amino acid sequences of LCDR1, LCDR2, and LCDR3 are shown in SEQ ID NO:4, 5, and 6, respectively. The antibody provided by this invention has good affinity and specificity for MSLN antigens, effectively detecting target proteins in cells overexpressing MSLN. It can also be widely used as a core ingredient in the development of high-sensitivity MSLN detection kits such as ELISA, Western blotting, and immunohistochemistry.

This invention relates to a method for constructing a polycystic kidney disease (PCD) model and its applications. The method involves overexpressing the MYCN gene in a target animal to obtain a PCD model that leads to PCD-related phenotypes. The method includes the following steps: obtaining a first strain of mice with Rosa26 knock-in overexpressing the CAG-LSL-HA tag-MYCN-IRES-BFP-Wpre-polyA gene; obtaining a second strain of mice by inserting Cre-WPRE-polyA into the start codon of the Pax8 gene; and crossing the first and second strains of mice to obtain a MYCN-overexpressing PCD model. This invention employs various experimental methods for validation, including histopathological analysis, immunohistochemical staining, and Western blotting. The model provided by this invention overcomes the limitations of existing in vitro cell models, organoids, and existing animal models in terms of limited phenotypes. The established MYCN-overexpressing PCD animal model exhibits stable disease progression and a short disease cycle, which not only helps to elucidate the disease mechanism but also serves as an ideal platform for drug screening and efficacy evaluation.

This invention discloses a monoclonal antibody against Oit3 protein in mouse hepatic sinusoidal endothelial cells or its antigen-binding fragment and its applications, belonging to the fields of biotechnology and medical immunology. The monoclonal antibody or its antigen-binding fragment comprises a heavy chain variable region and a light chain variable region. The heavy chain variable region contains HCDR1, HCDR2, and HCDR3 with amino acid sequences as shown in SEQ ID NO. 3~SEQ ID NO. 5; the light chain variable region contains LCDR1, LCDR2, and LCDR3 with amino acid sequences as shown in SEQ ID NO. 8~SEQ ID NO. 10. This monoclonal antibody and the recombinant fluorescent antibody exhibit high affinity and high specificity, and can be effectively applied to enzyme-linked immunosorbent assay (ELISA), Western blotting, and immunofluorescence detection. They can serve as important antibody tools for basic research on mouse hepatic sinusoidal endothelial cells and for exploring the mechanisms of liver diseases. Based on its variable region sequence, a recombinant fluorescent antibody, Oit3-scFv-GFP, was also constructed. This recombinant fluorescent antibody enables one-step direct immunofluorescence staining, providing a convenient tool for in situ visualization of mouse hepatic sinusoidal endothelial cells.

This invention discloses a monitoring and control device for protein immunoblotting gel electrophoresis experiments, comprising: a housing; a color mark sensor assembly, including a color mark sensor disposed on the front side of the housing; and a power control assembly, including a power input interface and a power output interface disposed on the housing, and a power on / off control module disposed within the housing. The power on / off control module is electrically connected to the color mark sensor and controls the on / off state of the power input interface and the power output interface according to the sensing signal of the color mark sensor. This invention uses a color mark sensor to determine color difference changes during the electrophoresis experiment, enabling easier and more accurate identification of the imprint position, high detection accuracy, and eliminating reliance on manual identification; moreover, after identification, the power supply to the electrophoresis apparatus can be promptly and automatically cut off using the power control assembly.

This invention relates to the field of cell protein blotting and fixation technology, specifically disclosing a convenient single-cell blotting aid tool. The tool includes a control unit and connected to the control unit a shaker cleaning mechanism, a heat dissipation mechanism, a multi-throughput gradient electrophoresis and fixation mechanism, and a harmonic acceleration sedimentation mechanism. The multi-throughput gradient electrophoresis and fixation mechanism is mounted on the shaker cleaning mechanism, and the heat dissipation mechanism is located inside the multi-throughput gradient electrophoresis and fixation mechanism. The multi-throughput gradient electrophoresis and fixation mechanism includes a first mounting base, a second mounting base, an electrophoresis tank, and a multi-throughput biochip. The first and second mounting bases are located at the upper ends of the shaker cleaning mechanism. The blotting aid tool of this invention provides a high-voltage electric field with harmonic characteristics to the upper and lower electrodes via a high-voltage power supply system. The direction of the electric field between the upper and lower electrodes can be switched according to the cell charge characteristics. Under the action of the high-voltage electric field with harmonic characteristics, charged cells rapidly settle into the micropores of the multi-throughput biochip.

This invention relates to the field of molecular biology, specifically to the application of PRAS40 protein as a therapeutic target for Alzheimer's disease. This invention combines Western blotting, computer-generated virtual docking, and surface plasmon resonance (SPR) techniques to confirm that PRAS40 protein is a key target influencing the pathological progression of AD. Experimental results show that specific intervention (elimination or reduction) of PRAS40 protein levels is sufficient to significantly improve cognitive deficits and pathological characteristics in AD model mice, providing a clear molecular target for subsequent screening and development of novel anti-AD drugs.

The application discloses an ELISA kit for detecting anti-human laminin gamma 1 IgA autoantibody and application thereof. The kit comprises an enzyme-labeled plate coated with human laminin gamma 1 recombinant protein. Preferably, the human laminin gamma 1 recombinant protein is deglycosylated human laminin gamma 1 recombinant protein. Further, the application also provides application of the kit in preparation of a reagent for diagnosing mucous membrane pemphigoid, in particular Laminin gamma 1 IgA type pemphigoid. The kit is the first kit for detecting human laminin gamma 1 IgA antibody by using the ELISA method, has the advantages of simple operation, short time consumption and low cost compared with traditional immunoblotting method, and adopts full-length deglycosylated laminin gamma 1 recombinant protein as an antigen, thereby solving the problem of incomplete antigen epitope coverage in the prior art, and early detection of diseases can be realized, early treatment is achieved, and the life quality of patients is improved.

This invention discloses an anti-ERBB2 monoclonal antibody, its preparation method, and its applications, belonging to the field of biomedical technology. This anti-ERBB2 monoclonal antibody uses the intracellular domain of the human ERBB2 protein as an immunogen and is obtained through animal immunization, B cell screening, gene cloning, and recombinant expression techniques. It can specifically recognize the human ERBB2 protein, exhibiting high affinity and high specificity, and can recognize both native and denatured ERBB2 proteins. This monoclonal antibody can be applied in immunohistochemistry, immunofluorescence, Western blotting (WB), flow cytometry, ELISA detection, and tumor targeted therapy. Experimental results show that this antibody has low background and no non-specific binding, accurately detecting ERBB2-overexpressing tumor cells and tissues, and has significant application value in the diagnosis, prognostic assessment, and targeted therapy of ERBB2-related diseases such as breast cancer.

This invention discloses a method and its application for predicting the trans-splicing activity of fragmented inpeptides. The method takes the amino acid sequence of the fragmented inpeptide as input, extracts 1280-dimensional residue-level sequence embedding features using the ESM-2 protein language model, and uses AlphaFold2 for high-precision three-dimensional structure prediction to obtain the spatial coordinates of the residues. A graph structure is constructed with residues as nodes and residue pairs with a Cα atomic distance of less than or equal to 8 Å as edges. Node features include sequence embedding and chain affiliation information, while edge features include Cα distance and cross-chain connection identifiers. The graph structure is input into the DeepIntein deep learning model based on a graph attention network, and the splicing activity prediction probability is output after multi-layer graph attention convolution, global pooling, and fully connected layers. The prediction throughput of this invention is more than three orders of magnitude higher than that of the traditional Western blotting method, breaking through the limitations of homology sequence exploration space.

The application discloses an E3 ubiquitin ligase FBXO22-mediated molecular glue degrader and application thereof, and relates to the technical field of medicinal chemistry. Based on the principle of targeted protein degradation, a series of E3 ubiquitin ligase FBXO22-mediated molecular glue degraders are designed and synthesized. The structure of the molecular glue degrader is shown in formula I or formula II: wherein R 1 and R 2 at least one contains alkylamine substitution. The application screens the activity of the compound by high-content screening technology and Western blotting technology, confirms that the molecular glue degrader can induce the degradation of bromodomain-containing protein 4 and other proteins in its family by E3 ubiquitin ligase FBXO22, thereby can hinder the relevant downstream pathway signals, has the activity of inducing cancer cell death and anti-fibrosis, and is expected to provide a new drug research direction for the treatment of bromodomain protein-related cancer and fibrosis diseases.

Abstract A method for detecting a biological marker in a sample, preferably a biological marker on a cancer cell, the method comprising: (i) contacting the sample with at least one DNA aptamer coupled to a first reagent and wherein the aptamer specifically binds to a first biological marker in the sample to form a first complex; (ii) contacting the first complex of (i) with a second binding agent that specifically binds to the first reagent such that a second complex is formed, wherein the second binding agent is coupled to a reporter molecule; (iii) contacting the second complex (ii) with a substrate for the reporter molecule of the at least one second binding agent; and (iv) detecting the substrate reaction by formation of a reaction product thereby detecting the biological marker in the sample. Biological markers include cytokeratin 8 (CK8), cytokeratin 18 (CK18), human epidermal growth factor receptor 2 (HER2) and epithelial cell adhesion molecule (EpCAM). Abstract 41 / 50 FIGURE 40 Frozen section preparation 4 Sample fixation + Wash (10 dips in PBS) DNA aptamer staining DAB Substrate DAB substrate HRP FAM Anti-FITC conjugated with HRP FAM DNA Aptamer conjugated with FAM HRP Frozen section slide / Touch imprint slide / Cytology slide / Direct smear slide Anti-FITC-HRP Wash (10 dips in PBS) DAB Wash (10 dips in PBS) Hematoxylin counter-staining Wash under tap water Cove rs lip DAB brown precipitate at the location of the aptamer B o min 1 min 15 sec 5 min 15 sec 5 min 15 sec 5 min 15 sec 10 sec 1 min 1 min A + Wash (10 dips in PBS) Total time for aptamer-DAB staining: —18 min 4 1 / 5 0 A B 1 0 m i n 1 m i n D A B s u b s t r a t e 1 5 s e c H R P A n t i - F I T C 5 m i n D N A a p t a m e r s t a i n i n g H R P 1 5 s e c 5 m i n 1 5 s e c D N A A p t a m e r c o n j u g a t e d w i t h F A M 5 m i n D A B W a s h ( 1 0 d i p s i n P B S ) F r o z e n s e c t i o n s l i d e / T o u c h i m p r i n t s l i d e / C y t o l o g y s l i d e / D i r e c t s m e a r s l i d e c o u n t e r - s t a i n i n g 1 m i n W a s h u n d e r t a p w a t e r 1 m i n C o v e r s l i p F I G U R E 4 020 25 28 36 33 19 D ec 2 02 5 1 9 D e c 2 0 2 5 A B 1 0 m i n F r o z e n s e c t i o n p r e p a r a t i o n 1 m i n S a m p l e f i x a t i o n D A B 2 0 2 5 2 8 3 6 3 3 s u b s t r a t e 1 5 s e c W a s h ( 1 0 d i p s i n P B S ) H R P A n t i - F I T C 5 m i n D N A a p t a m e r s t a i n i n g H R P 1 5 s e c 5 m i n A n t i - F I T C - H R P T o t a l t i m e f o r 1 5 s e c D N A A p t a m e r W a s h ( 1 0 d i p s i n P B S ) c o n j u g a t e d w i t h F A M 5 m i n D A B s t a i n i n g : ~ 1 8 m i n W a s h ( 1 0 d i p s i n P B S ) F r o z e n s e c t i o n s l i d e / T o u c h i m p r i n t s l i d e / 1 0 s e c H e m a t o x y l i n C y t o l o g y s l i d e / D i r e c t s m e a r s l i d e c o u n t e r - s t a i n i n g 1 m i n W a s h u n d e r t a p w a t e r 1 m i n C o v e r s l i p F I G U R E 4 0

This invention discloses a method for establishing GLI1 deacetylated or pseudoacylated mutants, relating to the field of molecular biology technology; comprising: S1: identifying GLI1 protein acetylation sites; S2: amplifying GLI1 deacetylated or pseudoacylated mutant gene fragments; S3: amplifying plasmid vectors; S4: ligating the GLI1 deacetylated or pseudoacylated gene fragments to the plasmid vectors; S5: verifying GLI1 deacetylated or pseudoacylated mutant protein expression using immunofluorescence and Western blot (WB). The method for establishing GLI1 deacetylated or pseudoacylated mutants of this invention can stably maintain the GLI1 deacetylated or pseudoacylated state.

The application belongs to the technical field of biological medicine, and relates to a BCLAF1 protein Ser564 site phosphorylation antigen peptide, a specific antibody and a preparation method and application thereof. The application identifies BCLAF1 protein Ser564 as a key phosphorylation site through SIK2 kinase screening, and designs and synthesizes a phosphorylation antigen peptide based on the site. After immunizing animals, a polyclonal antibody specifically recognizing BCLAF1 Ser564 site phosphorylation modification is successfully prepared. The antibody has high sensitivity and strong specificity, is suitable for various detection platforms such as immunohistochemistry and immunoblotting, and can be used for evaluating tumor immunotherapy efficacy. The application provides a key tool for analyzing BCLAF1 phosphorylation regulation mechanism, and lays a theoretical foundation for developing an immune combined therapy strategy targeting a SIK2-BCLAF1-CCL5 axis.

The application provides an antigen epitope polypeptide and a paralichthys dentatus BMP2 protein specific antibody prepared by the antigen epitope, designs an antigen epitope through bioinformatics analysis, adopts a segmented recombination expression strategy to obtain BMP2-N and BMP2-C recombination proteins in view of the low expression efficiency of full-length BMP2 protein in a prokaryotic system, and uses the recombination proteins as immunogens to prepare polyclonal antibodies. It is verified through immunoblotting and immunofluorescence that the prepared antibodies can specifically recognize the paralichthys dentatus BMP2 protein, have high affinity and good reaction activity, and effectively avoid the problems of false negative or low signal caused by insufficient cross reactivity of the antibodies. On this basis, a BMP2 immunological detection kit is constructed, precise positioning and expression analysis of the BMP2 protein in mucous membrane tissues such as skin, gill and intestinal tract of the paralichthys dentatus can be realized. The antibody and the kit provide a reliable tool for studying the role of BMP2 in fish mucosal immunity, and have important scientific research value and application prospect.

The application discloses a method for semi-quantitative analysis of expression level of secreted protein of human joint cartilage cells by using immunoblotting technology. The method comprises the following steps: 1) preparing a secreted protein culture solution; 2) collecting a sample of the secreted protein culture solution; 3) performing dialysis desalting treatment on the sample; 4) concentrating the secreted protein culture solution to prepare a powder; 5) re-dissolving the secreted protein culture solution powder and quantifying the protein; 6) separating the protein components in the re-dissolved culture solution; and 7) detecting the expression level of the secreted protein in the re-dissolved culture solution sample by using immunoblotting method. The low-temperature dialysis and vacuum concentration technology can significantly improve the concentration of the secreted protein in the human joint cartilage tissue culture solution sample under the condition of maintaining the integrity of the protein structure, and is suitable for detecting the chemical fluorescence signal of the secreted protein in the sample by using the immunoblotting technology. The application provides an important cell reaction detection tool for studying the pathogenesis of bone and joint diseases caused by degenerative changes of cartilage tissue and researching and developing biological small-molecule drugs for treating the diseases.

This invention provides an amino-PRX (Amino-PRX) vector as a nucleic acid / protein carrier. Amino-PRX freely provides amino groups to the sgRNA and acidic amino acids of Cas9, and CDs rotate and move only by mixing Amino-PRX and Cas9RNP (automatic molecular imprinting), thereby efficiently and easily forming a complex with Cas9RNP and achieving efficient delivery of Cas9RNP into cells. Furthermore, by optimizing the structure of the axon / cap connector and the amino / CD connector, the dynamics of Cas9RNP within cells can be precisely controlled, improving genome editing efficiency.

This invention belongs to the field of microbial isolation and purification technology, specifically relating to a method for preparing magnetic molecularly imprinted microspheres targeting marine bacteria and their applications. The microspheres use nano-Fe3O4 embedded with SiO2 as a substrate and polydopamine as the imprinting layer, forming a three-dimensional recognition cavity within the imprinting layer that complements the surface morphology and chemical composition of *Roseobacterium spp.* ISM strain. Specifically, nano-Fe3O4 is synthesized via co-precipitation, and a SiO2@Fe3O4 composite substrate is prepared via a sol-gel method. Dopamine self-polymerization imprinting is performed using an intact *Roseobacterium spp.* ISM strain as a template, and the microspheres are obtained after template removal. These microspheres exhibit superparamagnetism, structural stability, and good dispersibility. They can specifically adsorb and rapidly magnetically separate target bacteria from complex marine bacterial communities within 30 minutes. The purified target bacteria achieve a purity of 99.04%–100% without damaging the strain's activity, providing a high-quality bacterial strain foundation for marine sulfur cycle research and ecological restoration.

This invention relates to the field of laboratory automation technology, specifically to a method and system for adaptive optimization of protein immunoblotting incubation and elution parameters. It includes acquiring basic experimental data containing target protein molecular weight and antibody affinity; generating a joint optimization scheme for incubation parameters and multi-round elution parameters through a pre-set coupled kinetic model, wherein the model describes a quantitative mapping relationship between incubation conditions and subsequent elution difficulty; performing incubation operations and monitoring the signal on the membrane in real time, dynamically terminating incubation based on the signal slope; during elution operations, dividing the membrane surface into multiple regions of interest based on molecular weight and applying differentiated elution intensities, while simultaneously monitoring the waste liquid signal in real time to dynamically terminate elution; in multi-round detection scenarios, dynamically adjusting subsequent elution parameters based on the signal retention rate of the previous round, and adjusting the initial incubation conditions through a pre-distortion coefficient to compensate for signal attenuation in multiple rounds.

This invention discloses a method for determining the concentration of unfractionated heparin based on anti-Xa factor activity, the reagents used for determination, and the preparation method, relating to the field of biotechnology. The method comprises reagent R1 and reagent R2, wherein reagent R1 contains the following components: bovine Xa factor, Tris-HCl buffer, a temperature-sensitive composite stabilizer, sodium chloride, a magnesium salt at a concentration of 0.5-5 mmol / L, a heparin-specific nucleic acid aptamer at a concentration of 0.1-0.2 μmol / L, and gold nanoparticles at a concentration of 10-30 nmol / L. This invention effectively solves the problems of insufficient stability of the reagents and large batch-to-batch variations caused by the easy inactivation of active components such as Xa factor in existing technologies. The preparation process employs refined processes such as gradient heating, low-temperature plasma treatment, molecular imprinting, and microfluidic homogenization, providing dedicated protection for the active components and ensuring both reagent homogeneity and long-term stability.

This invention discloses an antigen for inducing polyclonal antibodies against pigeon circovirus Rep protein, comprising the amino acid sequence shown in SEQ ID NO.2, as well as the coding gene of the antigen, an expression vector for the coding gene, a primer pair for the coding gene, and the polyclonal antibody against pigeon circovirus Rep protein. The polyclonal antibody provided by this invention specifically reacts with pigeon circovirus in pigeon tissues, exhibiting good reactivity, and can be applied to molecular biology research related to the identification of pigeon circovirus, such as immunoblotting, immunohistochemistry, or immunofluorescence techniques.

The application provides a method for preparing fish skin collagen peptides with high antioxidant activity by DHPM and UIO-66 molecular imprinting, and relates to the technical fields of aquatic product deep processing and bioactive peptides. The method comprises the following steps: fish skin pretreatment, DHPM-induced collagen protein unfolding, endogenous enzyme autolysis, crude separation, selective target enrichment by using UIO-66 material with histidine as a molecular imprinting template, and drying collection. The application destroys the dense triple helix structure of collagen protein by using the physical field effect of dynamic high pressure microfluidization, exposes the enzyme cutting site, uses the endogenous enzyme of fish skin for autolysis, and then uses the histidine molecular imprinting UIO-66 material to accurately capture the high antioxidant activity peptide segment, so that the enzymolysis efficiency and the antioxidant activity of the product are significantly improved. The application does not need to add exogenous enzymes, and has the advantages of green process, low cost and high product purity. The obtained fish skin collagen peptides can be widely applied to the fields of functional food, cosmetics and medicine.