44 results about "Protein blotting" patented technology

The present invention relates to a process for preparing anthrax protective antigen protein from E. coli using fed batch culture. This process creates a constitutively expressing system for rapid, efficient, cost-effective and high-level production of anthrax PA from E. coli. The steps of the process involves, transforming E. coli DH5α cells with the recombinant constitutive expression plasmid containing the PA gene to obtain recombinant DH5α cells and testing the PA expression by lysis of said cells followed by denaturing gel electrophoresis and Western Blotting technique using PA antibodies. This is followed by fermentation and harvesting of the high cell density cells. The said cells are solubilized using 6-8 Molar Urea and separated by centrifugation. The urea denatured PA is isolated from said supernatant and purified and thereafter eluted.

The invention belongs to a preparation method and application of a bovine VCAM-1 polyclonal antibody. The method comprises the following steps of (1) RNA (ribonucleic acid) extraction; (2) reverse transcription PCR (polymerase chain reaction); (3) primer design and synthesis; (4) gene fragment amplification; (5) gene clone; (6) prokaryotic expression vector building; (7) expression and identification of DNA (deoxyribonucleic acid) recombinant plasmid in escherichia coli; (8) recombinant protein purification; (9) protein blotting; (10) recombinant protein dialysis and concentration determination; (11) immunizing antigen preparation; (12) immunizing and blood collection; (13) serum separation. The polyclonal antibody provided by the invention can be used for identifying the bovine recombinant VCAM-1 protein and natural recombinant VCAM-1 protein; the valence of the VCAM-1 polyclonal antibody is high; the specificity is high. The antibody particularly aims at the bovine natural VCAM-1 protein.

Nanocrystals having a ZnTe core and methods for making and using them to construct core-shell nanocrystals are described. These core-shell nanocrystals are highly stable and provide quantum yields and stability suitable for applications such as flow cytometry, cellular imaging, and protein blotting, medical imaging, and other applications where cadmium toxicity is an issue.

The invention discloses a non-closed protein blotting method for quickly detecting low-abundance proteins.The non-closed protein blotting method includes steps of (1), separating total proteins by the aid of gel in a gradient manner; (2), blotting the total proteins separated in the gradient manner on PVDF (polyvinylidene fluoride) films from the gel; (3), soaking the PVDF films with the blotted total proteins in 100% methanol; (4), drying the PVDF films treated by the 100% methanol; (5), carrying out color development detection on the target proteins in the dried PVDF films.The total proteins are extracted from samples.The non-closed protein blotting method for quickly detecting the low-abundance proteins has the advantages that the low-abundance protein detection efficiency can be improved, and practical, sensitive and speedy detection means can be provided.

The invention relates to novel application of emodin in the field of pharmacy, i.e. the application of emodin in preparing medicaments for treating P2X3 mediated neuropathic pains/nervous system diseases. In experiments, the emodin is observed to have an effect on inhibiting algesia behavioral reaction and is further observed to have an effect on inhibiting the mRNA and protein expression of nerve cell P2X3 receptors of rat dorsal root ganglia in a neuropathic pain model and rat trigeminal ganglion in a trigeminal neuralgia model by applying the technologies of immunohistochemistry, in-situ hybridization, RT-PCR (Reverse Transcription-Polymerase Chain Reaction), protein blotting and the like. The experiments prove that the mechanism of the effect of the emodin on inhibiting chronic pains (neuropathic pains) and pains transmitted by trigeminal ganglion nerve cells is to block algesia information transmission mediated by a primary sensory nerve cell P2X3 receptor. The invention provides a novel method for preventing and treating the pains mediated by the neuropathic pains and/or the trigeminal ganglion nerve cells; meanwhile, the invention shows that the emodin has the effect on influencing the P2X3 receptor, which is beneficial to medicinal application on preventing and treating nerve system diseases related to the P2X3 receptor.

The invention relates to a swing system of a protein blotting analyzer. The system comprises: a shaking table comprising a first rotating shaft arranged at the edge at one side of the shaking table; a base, wherein the shaking table is pivoted to the base through the first rotating table; and one or more reaction tanks arranged on the shaking table.

The invention discloses a method for raising expression of carbamoyl phosphate synthetase in HepGL liver cancer cells. The method comprises steps: CPS1 genome DNAs extracted from HepGL liver cancer cells are subjected to bisulfite treatment; the amplified CPS1 promoter area of the treated DNAs is subjected to methylation specific gene amplification; the amplified methylation sequences are subjected to sequencing; TALENs expression plasmids and non-methylation homologous plasmids which are homologous with the methylation loci are constructed according to base sequences around methylation loci, wherein, the homologous non-methylation sequence in the homologous plasmids is a mutation sequence wherein CG is mutated into CA; the TALENs expression plasmids and the homologous plasmids are subjected to transfection into HepGL liver cancer cells, HepGL liver cancer cells with transfection generation are screened, and subjected to Q-PCR and western blotting processing, and the expression amounts of mRNA of CPS1 and proteins in the cells are detected. The method can enhance the ammonia metabolism capability of HepGL liver cancer cells.

The invention provides a thermo-sensitive western blot sensing microsphere based on a quantum dot and a preparation method of same and belongs to the technical field of material science and bio-separation engineering. The method includes the steps of: 1) with a functional protein molecule as a template molecule, combining the functional protein molecule with CdTe quantum dots, which are co-modified with glutathione-thioglycollic acid (GSH-TGA), through electrostatic charge effect by means of amino groups and carboxyl groups, which are rich in the functional protein molecule; 2) in a non-oxygenenvironment, initiating polymerization by means of potassium persulfate (KPS), wherein thermo-sensitive isopropyl acrylamide (NIPAM) is used as a functional monomer and N,N-methylene bisacrylamide (MBA) is used as a crosslinker, so that through the free radical polymerization, the thermo-sensitive western blot sensing microsphere based on the quantum dot is produced. Through mechanism of electrontransfer between the functional protein and the quantum dot, fluorescent sensing of the microspheres to the template protein is achieved, and the synthesized western blot microspheres has performances such as thermo-sensitive response, fluorescent detection, etc.

The invention relates to a method for simultaneously rotating two films and marking two proteins, namely TRX and procaspase12. The method includes the steps of total protein extraction of cells, protein quantifying, gel electrophoresis, protein blotting, simultaneously marking of antibodies, X-ray film developing, and the like. The invention aims at building up a method which is characterized in that two PVDF films are used for simultaneously marking and detecting the two target proteins by using different antibodies after SDS-PAGE and protein blotting are carried out on the total protein in cells, meanwhile, the method is compared with the traditional method which has the steps of stripping films and then marking antibodies, and the accuracy, the reproducibility, the stability, and the like of the method are evaluated. The invention is simple and easy to carry out, can well represent the test result, and provides a method which is simpler and easier and saves time and samples, thereby having actual application value.

The invention relates to a new purpose of resveratrol in the pharmaceutical field, that is, an application of resveratrol in the preparation of medicines for treating P2X3-mediated chronic pain diseases. Experimental observation results show that resveratrol can inhibit pain related behaviors, and observation results of experiments applied with technologies of immunohistochemistry, in situ hybridization, RT-PCR, protein blotting and the like show that resveratrol can inhibit the mRNA expression and the protein expression of a P2X3 acceptor of the dorsal root ganglia from a neuropathic pain model rat. Experiments confirm that the mechanism of the inhibition effect of resveratrol to chronic pains is the antagonism of the primary sensory ganglion P2X3 acceptor-mediated pain information transmission. According to the invention, a new chronic pain (neuropathic pain) control method is ascertained, and simultaneously a case that resveratrol has effects of P2X3 acceptor antagonists is disclosed, so above observations of the invention are in favor of the application of resveratrol in the preparation of purine X3 acceptor (P2X3 acceptor) specificity antagonists.

The present invention relates to a fluorescent cell binding assay combining pre-labeling and Western blotting. Intact cells are incubated with pre-labelled binders preferably followed by SDS PAGE (sodium dodecylsulphate polyacrylamide gel) separation and Western blotting. More closely, the invention relates to a cell binding assay in which the degree or amount of binding of one or more cell interacting protein or protein component to the cell surface is measured with the ability to correlate the degree of cell binding to the sample load/total number of cells.

The invention discloses the novel use of a VEGF antibody and a VEGF receptor antagonist in the fields of pain-killing vaccines and pharmacy, namely the use of the VEGF antibody or the VEGF receptor antagonist in the preparation of an acute/chronic pain killing vaccine and a medicament for treating P2X3 and P2X2/3 purine receptor-mediated nerve system diseases. Through experiments it is observed that the VEGF antibody can inhibit the algetic behavior response; and by using immunohistochemistry, in-situ hybridization, RT-PCR, protein blotting and other techniques, it is observed that the VEGF antibody can inhibit the expression of mRNAs and proteins of the P2X2 and P2X3 receptors, VEGF and F1k-1/VEGFR2 of rat dorsal root ganglia of a model of neuropathic pain. Experiments prove that the pain killing mechanism of the VEGF antibody is that the pain is killed by neutralizing the VEGF or antagonist VEGF receptor to reduce pathologic pains and by blocking the pain information transmission of dorsal root neurons. The invention provides a new method for preventing and treating acute and chronic pains. Meanwhile, it is shown that the VEGF antibody can inhibit the expression of the P2X2 and P2X3 receptors, which is favorable for the use of the VEGF antibody and the VEGF receptor antagonist in the prevention and treatment of nerve system diseases related to the P2X2 and P2X3 receptors.

The invention belongs to the field of electrochemical biosensors, and particularly relates to a preparation method of an anti-pollution electrochemical biosensor based on temperature-sensitive westernblot gel. The preparation method comprises the following steps: polishing a glassy carbon electrode in an alumina suspension, carrying out ultrasonic treatment, and washing with ethanol and water insequence; respectively weighing acrylamide, N, N-methylene diacrylamide, ammonium persulfate, isopropyl acrylamide, transferrin or human immunoglobulin G and tetramethylethylenediamine, and uniformlydispersing in a phosphoric acid buffer solution to obtain a mixed solution; dropwise adding the prepared mixed solution to the surface of the pretreated GCE, carrying out free radical polymerization reaction in a nitrogen environment, and washing the surface of the electrode with water. The prepared biosensor is excellent in anti-pollution performance, and the anti-pollution performance is improved by 1-3 times; the detection sensitivity is high; the preparation method is simple and easy to operate.

The invention relates to a method for treating mitochondrial genetic diseases. The inventors have worked with primary fibroblasts from patients and control individuals and collected protein lysates for western blotting. Importantly, they observed that the genetic mitochondrial disorders, show a significant increase in phosphorylation of ribosomal protein S6 (pS6) compared to control fibroblasts, indicative of hyperactivated mTOR signaling. Patients with mitochondrial disorders and controls cells were treated for 48 hours with DMSO or BYL719. All lines from patients with mitochondrial diseases show reduced membrane potential, determined by TMRE staining intensity, and abnormal morphology, fragmentation and the presence of depolarized (low TMRE staining) mitochondria. Treatment with BYL719 attenuated these phenotypes in all MELAS fibroblasts while having no overt impact on the control cells. Similar experiments using flow cytometry confirmed membrane potential (TMRE) rescue by BYL719 treatment in MELAS fibroblasts.

The invention relates to the technical field of in-vitro detection, in particular to a detection method of a marker for diagnosing colorectal cancer. The detection method comprises the following steps of (1) extracting the total protein of cells or tissues; (2) preparing a separation gel and a concentrated gel; (3) performing polyacrylamide gel electrophoresis; (4) transferring a film; (5) incubating an antibody; and (5) developing by a chemiluminescence method. According to the present invention, the content of DCD in tissues or cells is detected through western-blot, so that whether the tissues or cells are intestinal cancer tissues or intestinal cancer cells or not is judged, DCD is a candidate diagnostic index and therapeutic target of colorectal cancer, and a new diagnostic thought can be provided for clinic.