131 results about "Western blot" patented technology

The invention discloses a construction and verification method of an HEK293T cell line capable of stably overexpressing human ANT2, which comprises the following steps: cloning a human ANT2 gene, constructing the human ANT2 gene into a lentivirus expression plasmid pLVX-TRE3G, co-transfecting the lentivirus expression plasmid pLVX-TRE3G and a helper plasmid pLVX-Tet3G into an HEK293T-ACE2 cell, and carrying out puromycin and G418 pressurized screening and Western blot identification to obtain the HEK293T cell line capable of stably overexpressing human ANT2. A result shows that the expression quantity of the ANT2 is the highest when the HEK293T-ACE2-ANT2 cell is induced for 12 hours at the Doxycycline of 6mg / L; when the ANT2 is induced to express, the virus nucleic acid load and the N protein expression level after SARS-CoV-2 infection are obviously reduced, which indicates that the ANT2 has an inhibition effect on SARS-CoV-2 infection replication, and a cell model and an experimental basis are provided for exploring the effect of the ANT2 in virus infection resistance.

The invention provides an immunofluorescence detection method and system for dynamically detecting mouse uterus structure development based on multiple time points, and immunofluorescence staining comprises the steps of sheet unfolding, dewaxing, repairing, cell membrane punching, sealing, primary antibody treatment and secondary antibody treatment. The system comprises a sample pretreatment control module, a multi-channel confocal microscope imaging module and a data analysis module integrating fluorescence intensity calculation, Pearson co-localization analysis and a Western blot data integration algorithm. Through a multi-time-point and multi-index immunofluorescence detection method, dynamic and systematic data support is provided for mouse uterus development mechanism research, and an application basis is provided for fundamental research of reproductive biology and construction of related disease models.

The invention relates to the technical field of biological medicines, and discloses a blocking antibody targeting CXCL6 and an application of the blocking antibody in preparation of antitumor drugs. The antibody or antigen-binding fragment thereof is capable of binding specifically to human CXCL6 protein with high affinity (e.g., Kd < = 1 nM). According to the invention, the specificity and high affinity of the antibody are verified by Western Blot, immunofluorescence, immunohistochemistry and surface plasmon resonance (SPR) technologies. Functional experiments show that the antibody can effectively block CXCL6-induced neutrophil chemotaxis, including in Transwell, a three-dimensional gel model and a zebra fish living body model. Besides, in a mouse MC38 colon cancer model, the antibody shows remarkable anti-tumor activity and can inhibit tumor growth driven by CXCL6. The invention also provides a pharmaceutical composition containing the antibody, and application of the antibody in preparation of drugs for preventing or treating diseases (such as inflammatory diseases, autoimmune diseases and cancers) related to abnormal expression or activity of CXCL6.

The present application relates to the technical field of biotechnology, in particular to a programmed death receptor ligand 2 monoclonal antibody and application thereof. The present application provides a PD-L2 monoclonal antibody, amino acid sequences of three CDR regions of a heavy chain thereof; amino acid sequences of three CDR regions of a light chain thereof; or amino acid sequences obtained by substitution, deletion or addition of one or more amino acids of the amino acid sequences, or amino acid sequences functionally identical or similar to the amino acid sequences; or amino acid sequences having at least 80% homology with the sequences. The present application widens the possibility of antibody therapy for blocking PD1 / PD1 ligand, and compared with existing PD-L2 monoclonal antibodies, the 8E3 antibody has unique amino acid sequences and CDR region sequences, high affinity and high specificity. The 8E3 antibody can be used in ELISA, Western blot and flow cytometry detection, and has wide application.

The invention discloses application of a PRTFDC1 gene in promoting porcine skeletal muscle satellite cell differentiation, myotube formation and skeletal muscle development. According to the invention, overexpression and knock-down experiments of the PRTFDC1 are respectively carried out in pig MuSCs, and immunofluorescence, qRT-PCR and Western blot analysis are carried out on cells subjected to induced differentiation, so that the PRTFDC1 is effectively shown to play a positive regulation role in a pig MuSCs differentiation process, and a new marker gene which can be applied to pig molecular breeding and growth performance evaluation is provided; and a novel molecular target and a novel detection tool are provided for muscle development regulation and biological breeding.

The invention relates to the field of antibodies, and particularly provides a high-affinity PADI2 antibody and a preparation method thereof. According to the invention, eight high-affinity PADI2 antibodies are obtained by screening through an antibody pair screening experiment, and the antibodies can be specifically combined with PADI2 protein and have high affinity. Experimental results show that the antibody disclosed by the invention shows excellent binding capacity in ELISA (Enzyme-Linked Immunosorbent Assay), Western blot and cell experiments, and can be used for detecting, diagnosing and treating PADI2 related diseases.

The invention provides an application of 2-indolecarboxylic acid in preparation of a tumor targeted therapy drug. Through computer-aided drug design and molecular docking optimization, 2-indolecarboxylic acid shows high binding energy to an HER2 kinase structural domain. In HER2 positive NCI-N87 gastric cancer cells, the 2-indolecarboxylic acid significantly inhibits cell proliferation and induces G0 / G1 phase arrest and apoptosis. The mechanism research shows that the 2-indoleformic acid can play a role by regulating and controlling the HER2 / Akt / beta-catenin pathway (Western blot verification). In an NCI-N87 cell tumor-bearing nude mouse model, 2-indolecarboxylic acid (15 mg / kg) significantly inhibits tumor growth, and does not cause obvious weight loss or organ toxicity. Immunohistochemical analysis shows that the positive rate of a tumor tissue proliferation marker Ki-67 in a treatment group is remarkably reduced, and apoptosis detection shows that the proportion of TUNEL positive cells is remarkably increased, which indicates that 2-indolecarboxylic acid inhibits tumor cell proliferation and promotes apoptosis at the same time. The invention provides a lead compound with clinical transformation potential for HER2 targeted therapy.

The invention discloses application of ailanone in preparation of a medicine for treating feline infectious peritonitis, and belongs to the field of biological medicine. The change of the expression quantity of the feline infectious peritonitis virus N protein is detected through indirect immunofluorescence and western blot, and the change of the quantity titer of the feline infectious peritonitis virus is detected through TCID50. Results show that ailanone has an obvious effect of resisting the feline infectious peritonitis virus, the inhibitory effect on the feline infectious peritonitis virus is enhanced along with increase of the drug concentration, and the half effective concentration is 0.09 mu M. Meanwhile, drug inhibition tests show that ailanone has no inhibition effect on feline calicivirus. The invention provides a novel therapeutic drug and strategy for treating feline infectious peritonitis.

The invention discloses a down-regulated CDC20-based breast cancer cell ferroptosis sensitivity detection method, and belongs to the technical field of detection methods, and the method comprises the following steps: cell grouping and transfection: culturing a breast cancer cell system, and setting an experimental group and a transfection negative control group; wherein in the experimental group, CDC20-siRNA transfection is carried out on breast cancer cells; an experimental group with the CDC20 relative expression quantity lower than 0.5 is screened through Western blot to serve as an optimal experimental group, and subsequent experiments are conducted; and then ferroptosis induction and ferroptosis induction and inhibition are carried out, and related detection is carried out. According to the invention, CDC20 is used as a regulation target of breast cancer ferroptosis sensitivity for the first time, and the target limitation of a traditional method is broken through; and the reliability and repeatability of a detection result are ensured through a standardized operation process and closed-loop verification logic.

The invention belongs to the field of pharmacy, and relates to a protein degradation targeting chimera (PROTAC) taking PD-L1 as a target spot and medical application thereof. The protein degradation targeting chimera is a compound composed of a target protein ligand, an E3 ligase ligand and a linker and a pharmaceutically acceptable salt of the compound, the specific structure of the compound is as shown in a formula I, and substituent groups are described in the specification in detail. A Western blot experiment (Western Blot) shows that the compound disclosed by the invention can be used for degrading the PD-L1 protein in a targeted manner.

The invention discloses a method for improving autism behaviors through Npas4 expression and overexpression in a cat chat syndrome model, and belongs to the technical field of animal model construction. Selecting SD wild rats and CdCS model rats, and normally feeding and breeding the rats; the method comprises the following steps: measuring the protein concentration by adopting a BCA method, then carrying out Western Blot, then carrying out brain stereotactic injection, and finally carrying out behavioral testing. According to the invention, a CdCS rat model is established by using a CRISPR-Cas9 technology, and the model shows cognitive and social behavior disorders and medial prefrontal cortex (mPFC) neuron dendritic abnormality; in addition, an abnormally expressed gene is found in the model mouse, the research focuses on Npas4, and a new insight is possibly provided for disease treatment.

This invention belongs to the field of traditional Chinese medicine treatment of acute kidney injury, and provides a traditional Chinese medicine composition for treating sepsis-related acute kidney injury and its preparation method. It is prepared by mixing honeysuckle, forsythia, roasted peach kernel, raw rhubarb, raw astragalus, wine-processed angelica, cinnamon twig, mirabilite, and raw licorice in a certain proportion. It detoxifies and removes blood stasis, invigorates qi and unblocks the meridians. It is used for persistent high fever and difficulty in urination and defecation caused by internal accumulation of toxic heat and blood stasis. Symptoms include oliguria or dribbling, red tongue, and rapid pulse; it is also used for sepsis patients with the above symptoms. Treatment with the detoxifying and blood-stasis-removing granules improved survival rate, effectively restored some renal function, reduced renal tubular damage, and decreased TNF-α and IL-1β expression. Immunohistochemical and Western blot results were consistent. The detoxifying and blood-stasis-removing granules can alleviate sepsis-induced acute kidney injury.

The utility model discloses Western blot full-automatic multi-round circulating detection equipment, which realizes continuous detection of multi-target proteins on a single transfer printing film by integrating a liquid treatment module, an incubation reaction module, a signal capture module and a program control module. Residues are removed by adopting antibody denaturation liquid, signal crosstalk is avoided by combining a chemiluminescence system, 20 cycles of cyclic operation are supported, and a self-cleaning function is achieved. Equipment is compatible with a conventional transfer printing film, and the detection flux and the standardization level are remarkably improved.

The invention belongs to the technical field of biological genes, and discloses a method for identifying RNA interaction protein based on a Turbo ID technology and an MCP / MS2 system, the method can be used for identifying protein interacting with RNA in vitro or in vivo, and the method specifically comprises the following steps: firstly, respectively constructing RNA expression vectors inserted with MS2 tags, then constructing fusion protein vectors expressing MCP-Turbo ID, and finally constructing fusion protein vectors for expressing MCP-Turbo ID; the method comprises the following steps of: inserting RNA (Ribonucleic Acid) into an MS2 tag, transgenosis into a plant receptor by utilizing an agrobacterium tumefaciens mediated genetic transformation method to obtain a transgenic plant material expressing MCP-Turbo ID, combining the RNA inserted with the MS2 tag with the MCP-Turbo ID in vivo / in vitro by utilizing a hybridization or in-vitro incubation method, further enabling the Turbo ID to be adjacent to the RNA, biotinylating the RNA binding protein, and obtaining a transgenic plant material expressing the MCP-Turbo ID. And then enriching biotinylated protein by adopting streptavidin magnetic beads, and identifying and analyzing through Western Blot and mass spectrometry. The method not only solves the problem that the interaction protein of specific RNA in a living cell environment cannot be effectively identified in the prior art, but also can capture a transient or low-affinity RNA-protein interaction relationship.

The invention relates to the technical field of genetic engineering, in particular to a recombinant pichia pastoris strain for expressing six tandem antibacterial peptides MPX as well as a construction method and application of the recombinant pichia pastoris strain. Aiming at the technical problems that the expression quantity of MPX in pichia pastoris is low, MPX is easily degraded by endogenous protease, the product activity is insufficient and the like, the invention constructs a recombinant pichia pastoris strain for expressing the six-tandem antibacterial peptide MPX, and a construction method comprises the following steps: designing and constructing a coding sequence of the six-tandem antibacterial peptide MPX; and a recombinant expression vector pPICZ alphaA-6MPX for expressing the six-tandem antibacterial peptide MPX is constructed. A linearized recombinant vector is introduced into pichia pastoris GS115 through a chemical conversion method, a positive transformant is screened through a bleomycin-containing selective culture medium, and a recombinant strain for stably expressing six tandem MPX is obtained by combining PCR and Westernblot identification. The recombinant strain can be used for inhibiting growth of pathogenic bacteria and has application potential as an antibacterial active preparation.

The present application relates to the field of antibodies, and in particular, the present application provides a high affinity PADI2 antibody and a preparation method thereof. Through antibody screening experiments, eight high affinity PADI2 antibodies are screened, the antibodies can specifically bind to PADI2 protein, and have high affinity. Experimental results show that the antibodies of the present application exhibit excellent binding capacity in ELISA, Western blot and cell experiments, and can be used for detection, diagnosis and treatment of PADI2 related diseases.

The invention relates to an anti-ARDS cationic liposome delivery system and application thereof, in particular to an anti-ARDS cationic liposome delivery system and application thereof. The sivelestat sodium carrier is composed of DOTAP, DSPC, polyethylene glycol and cholesterol, and can load two micro RNAs (miR125 and miR223) and a fat-soluble drug sivelestat sodium at the same time. Through TEM (transmission electron microscope detection), Western Blot, real-time fluorescent quantitative PCR and LPS-induced mouse ARDS model pharmacological experiments, it is proved that the material can play a better anti-inflammatory role, the treatment effect is remarkable, and the material has a very good development prospect.

The invention provides a traditional Chinese medicine composition, a traditional Chinese medicine preparation, a preparation method and application, and particularly belongs to the technical field of traditional Chinese medicine preparation. The traditional Chinese medicine composition comprises the following components in parts by weight: 7-11 parts of champignon, 19-29 parts of aloe, 10-14 parts of fructus chebulae immaturus, 5-7 parts of rhizoma zingiberis, 5-7 parts of rhizoma nardostachyos, 4-6 parts of fructus chebulae and 2-4 parts of halite. The traditional Chinese medicine composition can improve the activity of SOD, inhibit the content of MDA, reduce damage of oxygen free radicals to gastric mucosa, effectively relieve inflammatory response, achieve anti-inflammation, improve ulcer surface blood flow, improve gastric mucosa damage, reduce ulcer surfaces and promote ulcer healing, and therefore the protective effect on gastric ulcer is achieved. The results of network pharmacology, molecular docking speculation and Western blot method verification show that the FMST plays an anti-inflammatory role through a TLR4 / MyD88 / NF-kappa B signal channel.

The invention relates to the technical field of biological medicines, and particularly discloses an anti-PRRX2 monoclonal antibody and application thereof in detection of gemcitabine drug resistance of adenocarcinoma. The antibody is prepared through a hybridoma technology, a BALB / c mouse is immunized with a recombinant human PRRX2 protein, and a high-affinity antibody strain is obtained through cell fusion, screening and monocloning. Sequences of light and heavy chain variable regions of the antibody are shown as SEQ ID No: 7 and SEQ ID No: 8, and sequences of a CDR region are shown as SEQ ID No: 1-6. After the antibody is coupled with HRP, the antibody is used for Western Blot to detect PRRX2 expression in a pancreatic ductal adenocarcinoma (PDAC) patient sample, and the result shows that the detection sensitivity to a gemcitabine drug-resistant sample reaches 96.6%, and the specificity is 100%. The antibody disclosed by the invention has high specificity and stability, provides a reliable tool for clinical drug resistance detection, and is beneficial to formulating an individualized treatment strategy.

This invention discloses the use of Chek2 inhibitors in the preparation of drugs for the prevention and / or treatment of postoperative pain sensitization induced or aggravated by chronic sleep disorders, and its intervention methods, relating to the field of biomedical technology. This invention provides the use of Chek2 inhibitors in the preparation of drugs for the prevention and / or treatment of postoperative pain sensitization induced or aggravated by chronic sleep disorders. This invention is the first to discover and validate Chek2 as a key molecular target linking "chronic sleep disorders" and "postoperative pain sensitization." By constructing a composite animal model of chronic sleep deprivation (CSD) combined with postoperative incision pain, the pathological process of increased postoperative pain in patients with sleep disorders was simulated. Using mRNA-seq and Western blot techniques, this invention is the first to discover and confirm that the expression level of Chek2 in the nucleus accumbens (NAc) brain region is significantly upregulated during postoperative pain sensitization induced by chronic sleep disorders, filling a gap in the field of "sleep disorder-pain interaction" regarding specific central targets.

The invention belongs to the technical field of gene detection, and particularly relates to a brand new application of a DDX42 gene or an expression product thereof as an acute myelogenous leukemia (AML) diagnosis and prognosis biomarker. Specific high expression of DDX42 in bone marrow and peripheral blood samples of AML patients is revealed for the first time, the expression level of DDX42 is remarkably different from that of a healthy control group, and DDX42 can be detected through real-time fluorescent quantitative PCR, western blot and other methods and is used for diagnosis, differential diagnosis, disease state evaluation or risk stratification of AML. Meanwhile, the invention clarifies a molecular mechanism that DDX42 promotes AML cell proliferation and inhibits apoptosis and differentiation through PLK1 and LIMK1-CREB-DNMT1 signal axes, and verifies that a transcription factor ERG is an upstream forward regulatory factor for the first time. Based on the discovery, the invention not only provides a novel high-sensitivity molecular diagnosis marker for AML, but also lays a solid experimental foundation for developing a novel AML treatment strategy and a drug screening system targeting DDX42 and related signal channels thereof. The method has the advantages of mature detection method, high marker relevance, clear application prospect and the like.

A method and system are disclosed for differentiating non-human primate induced pluripotent stem cells (NHP iPSCs) into hepatocyte-like cells through a defined, multi-stage protocol. The method includes sequentially exposing NHP iPSCs to six distinct cell differentiation media over an approximate eighteen-day period. Each medium comprises defined combinations of growth factors, small molecules, and inhibitors that direct progressive lineage specification through anterior primitive streak, definitive endoderm, posterior foregut, liver bud progenitors, hepatoblasts, and hepatocyte-like cells. Concentration ranges and exposure durations are provided for each stage, ensuring reproducibility and reducing heterogeneity across cell populations. In some embodiments, treatment with a transforming growth factor beta (TGF-β ) receptor I inhibitor during early hepatoblast specification enhances lineage fidelity. The resulting hepatocyte-like cells are validated by immunostaining and western blotting for hepatic markers. Applications include preclinical drug metabolism, hepatotoxicity assessment, and in vitro disease modeling using non-human primate–derived systems.

The invention relates to the technical field of biological medicines, in particular to application of wild buckwheat rhizome freeze-dried powder in preparation of a medicine for preventing or treating antimony pneumoconiosis. A mouse antimony pneumoconiosis model is constructed by injecting an antimony (Sb) suspension into a trachea, and pulmonary fibrosis lesion caused by antimony exposure can be remarkably relieved by intragastric administration of the wild buckwheat rhizome freeze-dried powder prepared by the invention. Experimental results show that Hamp; e dyeing indicates that the freeze-dried powder can obviously relieve the fibrosis severity of the lung tissue; western Blot and PCR (Polymerase Chain Reaction) detection results show that the wild buckwheat rhizome can partially inhibit the expression up-regulation of Col-1 and alpha-SMA induced by antimony in protein and gene levels; in addition, the wild buckwheat rhizome can also obviously reduce the content of hydroxyproline in lung tissues. The results show that the wild buckwheat rhizome freeze-dried powder can effectively improve multiple pathological indexes and reduce the pulmonary fibrosis degree, has good potential in the aspect of preventing antimony pneumoconiosis, and has potential application value.

The application relates to an active pulmonary tuberculosis marker and application thereof, the active pulmonary tuberculosis marker is PCBP1, the expression of the PCBP1 in peripheral blood mononuclear cells of an active pulmonary tuberculosis patient is obviously down-regulated, the down-regulated PCBP1 has certain significance for diagnosis of the active pulmonary tuberculosis patient. Moreover, the expression amount of the PCBP1 in the APTB patient is detected by using an RT-qPCR and western blot method, the disease can be diagnosed, and then the active pulmonary tuberculosis diagnosis reagent can be prepared. The application discloses that the PCBP1 can be used as the active pulmonary tuberculosis marker for the first time, and provides a new direction for clinical diagnosis of the active pulmonary tuberculosis.

The invention discloses an application of Lycobetaine in the preparation of an SHMT2 protein targeted inhibitor. According to the invention, 11 candidate compounds are obtained by adopting a molecular docking technology, and a CCK-8 cell viability experiment shows that the anti-proliferative activity of Lycobetaine on non-small cell lung cancer A549 and H1299 cells is superior to that of a standard SHMT2 inhibitor SHIN1. A Western blot result shows that the expression of the SHMT2 is down-regulated by the Lycobetaine in a dose-dependent manner, and the SHMT1 is not influenced. A gene knockout experiment shows that the anti-proliferative activity of the Lycobetaine depends on the expression of the SHMT2. In-vitro experiments and organoid models show that Lycobetaine significantly inhibits tumor growth and induces structural collapse and volume reduction of tumor organoids. The Lycobetaine can be used as an SHMT2 protein targeted inhibitor, and is applied to tumor treatment.

The utility model relates to the technical field of molecular biology experimental equipment, in particular to a locating and fast-assembling Western Blot film transferring clamp system which comprises a film transferring clamp base body, a buckle connecting piece capable of being fast connected is installed between the opening ends of the film transferring clamp base body, a sponge layer is arranged in the film transferring clamp base body, and a locating piece is arranged on the sponge layer. A first pressure-sensitive adhesive tape is bonded between the corner of the bottom of the sponge layer and the bottom of the transfer film clamp base body, filter paper is placed on the top of the sponge layer, a second pressure-sensitive adhesive tape is bonded between the corner of the top of the sponge layer and the bottom of the filter paper, and a polyvinylidene fluoride film is placed above the filter paper. Through a bonding, locking and scale cooperative positioning mechanism, the membrane transfer success rate and the experiment repeatability are remarkably improved, meanwhile, the operation complexity is reduced, and a reliable tool is provided for standardization of the western blot technology.

The invention discloses a targeted protein degradation agent utilizing a ubiquitin-proteasome pathway as well as a preparation method and application of the targeted protein degradation agent, and belongs to the technical field of biological medicines. On the basis of a PROTAC (protein targeted degradation chimera) strategy and on the basis of imatinib, different Linkers are introduced to be connected with an E3 ubiquitin enzyme ligand Nutlin-3 derivative, a degradation tag is labeled on BCR-ABL cancer protein, and the degradation agent with the Bcr-Abl protein targeting capacity is constructed. A Western blot experiment shows that the targeted protein degradation agent shows a good degradation effect on the Bcr-Abl protein in K562 cells, and the compound WP shows a degradation effect on concentration dependence and time dependence of the Bcr-Abl protein. Tumor cell proliferation experiments show that the compound has certain inhibitory activity on K562 cells. Wherein when the Linker is dodecane-1, 12-diketone, the anti-proliferative activity is the best. An apoptosis experiment and a period experiment show that the targeted protein degradation agent can realize concentration-dependent promotion of cell apoptosis and retard cells in S and G1 / G0 periods.

The invention relates to the field of antibodies, and particularly provides a high-affinity PADI2 antibody and a preparation method thereof. According to the invention, eight high-affinity PADI2 antibodies are obtained by screening through an antibody pair screening experiment, and the antibodies can be specifically combined with PADI2 protein and have high affinity. Experimental results show that the antibody disclosed by the invention shows excellent binding capacity in ELISA (Enzyme-Linked Immunosorbent Assay), Western blot and cell experiments, and can be used for detecting, diagnosing and treating PADI2 related diseases.

The invention relates to the field of biological medicine, in particular to a lactic dehydrogenase LDHB protein 196-site serine phosphorylation antibody as well as a preparation method and application thereof. On the basis of the 196-site serine phosphorylated polypeptide of the lactic dehydrogenase LDHB protein, the 196-site serine phosphorylated antigen and antibody of the lactic dehydrogenase LDHB protein are prepared, and a preparation method of the 196-site serine phosphorylated antibody of the lactic dehydrogenase LDHB protein is provided. The prepared lactic dehydrogenase LDHB protein 196-site serine phosphorylated antibody can be used for detecting low-abundance LDHB Ser196 signals, is suitable for various detection platforms such as Western blot and immunohistochemistry, and meets the detection requirements of tumor tissue samples.