66 results about "Protein spot" patented technology

The invention discloses a Coomassie Brilliant Blue G250 staining liquid, which contains the following components: CBB G-250 with a mass volume concentration of 0.1-0.2%, ammonium sulfate with a mass volume concentration of 5-15%, and a volume percentage of 3 -8% phosphoric acid, 20-40% ethanol by volume, and 5-20% methanol by volume. The staining solution integrates the fixative solution, the sensitizing solution and the staining solution, the background of the dyed gel is lighter, and the dyeing sensitivity is high; a method for dyeing by using the above-mentioned Coomassie Brilliant Blue G250 staining solution is also disclosed. In addition to high repeatability and low staining background, this detection method is fast and sensitive, and the number of detected protein spots is relatively large; it has good mass spectrometry compatibility and can reduce protein modification in vitro; it has a wide range of applications. At the same time, the application of the above method in the qualitative and/or quantitative detection of proteins is disclosed.

The invention belongs to the field of biological chemistry and specifically relates to extraction method of chloroplast protein of xylophyta ginkgo leaf. The method comprises the following steps of: firstly, preparing coarse chloroplast particles from the ginkgo leaves, performing gradient centrifugation through Percoll, so as to obtain complete chloroplast by a specific centrifugal force; then, dissolving protein by using a protein extraction buffer solution, adding the same volume of Tris-balanced phenol for extraction, and precipitating protein by using a ammonium acetate methanol solution. The extraction method is capable of effectively increase extraction efficiency and purity of chloroplast protein. The ginkgo leaf chloroplast protein obtained by the method can be used for proteomics analysis of ginkgo leaf and other xylophyta materials rich in interfering substances, so that solubility and purity of protein are improved; a 2-DE map of the protein has a clear background and many protein spots are distinguishable.

The invention discloses a method for obtaining a wheat root related drought resistant protein through utilizing dimensional electrophoresis and an MALDI-TOF-MS technology. The method comprises a step of material treatment, a step of dimensional electrophoresis, a step of atlas analysis, and a step of MALDI-TOF-MS, and the MALDI-TOF-MS step is characterized in that differential protein spots undergo in-gel enzymatic hydrolysis, ultraflex TOF/TOF is used to carry out MALDI-TOF mass spectrum fingerprint analysis, and obtained PMF data are identified through retrieving Protein Data Bank by Mascot software. In the invention, the combination of dimensional electrophoresis and the MALDI-TOF-MS technology is adopted, the protein expression difference between the drought stressed wheat root and the normal-water-supply wheat root is analyzed, and the identification is further carried out through the mass spectrum, so the root expression protein obtained through the method is directly linked to the property, and is of greater significance than the gene level and the other organs of crops.

The present inventions is directed to a method and computer program product for detecting and quantifying protein spots, including: generating an average gel image by taking a pixel-by-pixel average of the intensities of a plurality of aligned gel images; detecting spots on the average gel image using pinnacle detection; and quantifying spots on individual gels using the maximum intensity within fixed neighborhoods surrounding pinnacle locations found in the average gel image.

The invention discloses a method for screening and identifying an ECH1 autoantibody and an application thereof. The method comprises the following steps: adopting a lung cancer cell line H1299 holoprotein extractive for performing two-dimensional electrophoresis; adopting a western blot method for processing the serum of 5 patients with lung cancer and serum of 5 normal people, screening the significant tumor autoantibody, and after comparative analysis, only further analyzing the protein spots appearing in the serum of the patients; cutting and digesting the interesting protein spots on SDS-PAGE gelatin, and then adopting a LG-MS/MS mass spectrometry method for analyzing the protein spots, detecting and identifying as ECH1; and further verifying if the ECH1 autoantibody can be used as a lung cancer diagnosis marker. Through research, the invention finds that the detection for the serum ECH1 autoantibody according to an ELISA method can be used for more accurately identifying the patients with lung cancer from the normal people, identifying the patients with chronic obstructive pulmonary diseases and early diagnosing the lung cancer.

After the sequencing of the human genome, great interest has developed in trying to discern the complementary proteome of humans and other species. The present disclosure provides devices, systems, and methods for proteomic fractionation that may increase the number of protein spots visualized by 2DE analysis, and may allow enrichment of proteins normally not detectable by standard 2DE analysis. According to some embodiments of the disclosure, devices, systems, and methods of the disclosure relate to fractionating a proteome on the basis of surface charge, hydrophobicity, isoelectric point and/or size.

The present invention relates to a method and device for the removal of fluids and vapors in containment, from adjacent surfaces including supporting flat surfaces and substrates, such as protein spots printed in arrays and micro-array formats. This method and device removes fluids and vapors under controlled and repeatable conditions, enabling uniform phases for degree of drying incurred by objects, specimen and structures, including tissues, adsorbed particles and biological substrates, while reducing and preventing impact of meniscus phase surface tension forces.

The invention discloses an acquisition method for extraction of total lily protein and a two-directional electrophoresis differential protein map. In the method, the rotating speed is set to be 20000 rpm in the extraction of total lily protein, and the total lily protein is precipitated and centrifuged at least three times repeatedly, so that the extracted total lily protein is relatively white in color and good in quality; the mass detection step of the protein is added, and the mass of the extracted protein is detected in a sodium dodecyl sulfate polyacrylamide gel manner, so that the problem that the subsequent first direction isoelectric focusing effect is poor is avoided. The focusing program in the isoelectric focusing effect is improved, the time of two desalting steps such as step S1 and step S2 in the focusing program is prolonged to be 2.5h, and the salt of the protein is well removed, so that the focusing program is well completed. The two-directional electrophoresis differential protein map of the total lily protein obtained by the method is clear in background, more in protein spots and uniform in protein spot distribution.

A precast slab gel for use in submerged ("submarine") horizontal electrophoresis is formed in a tray that includes a flat base and two raised walls on opposing sides of the base, the walls containing one or more tabs on their outer surfaces, the tabs mating with grooves in the interior walls of the tank. The mating of the tabs with the grooves prevents movement and floating of the tray within to the electrophoresis cell during use. The tabs are designed to mate with grooves that are present in the tank for other purposes, which adds to the versatility of the design. Further versatility is achieved by joining the tabs to the tray walls by thin webs, which make the tabs readily removable, thereby rendering the tray usable in cell tanks that do not contain grooves. Further aspects of the invention include pins or posts extending upward from the tray base to anchor the gel, and the printing of indicia on the tray base by hot foil stamping, with the discovery that indicia printed in this manner are capable of producing a fluorescent image as part of the image produced by fluorescent-dyed protein spots in the electropherogram.

The invention belongs to the technical field of biology, and discloses an electrophoresis method for removing Rubisco from plant leaves and enriching and separating residual proteins. The method comprises the following steps of: removing Rubisco from watermelon leaves by using 18 percent PEG-4000; leaching residual proteins through a TCA-acetone precipitation method; and performing two-dimensional electrophoresis to separate residual proteins from watermelon leaves. Compared with a holoprotein two-dimensional electrophoretogram, the method has the advantages that: after Rubisco in watermelon leaves is precipitated by adopting the 18 percent PEG, most large and small subunits of Rubisco disappear, the expression levels of most residual proteins are raised, the quantity of proteins obtained by separating is increased by 48.4 percent, and a large quantity of new protein spots (65+/-15) appear in a region of which the molecular weight is smaller than 25.0kDa. As proved by repeated experiments, the electrophoresis method has the advantages of high repeatability, clear spectra and reliable result.

The invention discloses a gel protein partitioning method based on fuzzy clustering. The method comprises the steps that first, images are filtered, and the contrast ratio of the images is enhanced; second, the number of clustering categories, weighted indexes, an iteration termination threshold value, the maximum number of iterations and an initial clustering center are initialized; third, a radial width value in a kernel function is calculated; fourth, a membership matrix and a clustering center are updated; fifth, whether an absolute difference value of the current new clustering center and the last clustering center is smaller than the iteration termination threshold value or whether a current value of an iteration counter is greater than the maximum number of iterations is judged through comparison, if yes, the process is stopped, a final membership matrix and a final clustering center are output, and the sixth step continues to be executed, or else the fourth step continues to be executed after the iteration counter adds one; sixth, defuzzification is performed to obtain an optimal partitioning result. Through the method, noise eliminating ability is improved, relatively weak protein spots can be separated, and consequently more protein spots are separated; moreover, the method has a good separation effect on lightly-overlapping protein spots and has high partitioning precision.

The invention relates to a method for early diagnosing an acute kidney injury caused by a contrast medium by utilizing urine MBL (mannan-binding lectin), which comprises the following steps of: comparing fluorescence difference electrophoresis charts of the urine of a patient 24 hours after an operation and the basis urine of the patient before the operation, and searching out a plurality of differentially expressed protein spots; identifying through a matrix-supported laser desorption ionization time of flight mass spectrometry technique to discover that the mannan-binding lectin (MBL) and serine protease related to the MBL are both increased in the differentially expressed protein spots, and the difference has a statistical significance for prompting that the acute kidney injury caused by the contrast medium is related to a pathway of MBL (mannan-binding lectin) complement activation, which is verified by a patient with the kidney injury caused by the contrast medium through an urine ELISA (enzyme-linked immuno sorbent assay) way. If applied to the clinic, the method of the invention can be used for detecting the urine MBL (mannan-binding lectin) 24 hours after an operation through the ELISA (enzyme-linked immuno sorbent assay) way and early diagnosing the kidney injury caused by the contrast medium, which is 24 hours earlier than the diagnosis of serum creatinine, and provides basis for early clinical diagnosis, early treatment and improvement of prognosis.

The invention discloses an efficient extraction method for proteins in tea leaves. Fresh tea leaves are picked, a mixed TCA/acetone solution is adopted for extraction, separation and standing are conducted, sediments after standing are further washed and purified, low-temperature vacuum freezing drying is conducted, crude proteins are obtained, then lysate is adopted for splitting decomposition ofthe crude proteins, supernatant is obtained through centrifugation, and a pure protein solution is obtained. According to the method, the quantity of the proteins obtained through extraction is effectively increased, after 2D-PAGE is adopted for separation, the separation effect is good, the proteins can be effectively separated, and the protein spots are clear; LC/MS/MS is adopted for identifying the proteins, and the result shows that there are 4,117-4,218 proteins extracted through the method. The method lays an important foundation for the research of tea leaf proteomics.

The invention relates to 5 identified protein biomarkers, gamma- and beta-Actin proteins, for screening, diagnosis, drug targeting, and drug design for resistance of cancer to an Ab1 kinase inhibitor. The method is based on the use of two-dimensional (2D) gel electrophoresis to separate the complex mixture of proteins found in bone marrow aspirate samples, taken from patients at time of diagnosis of Chronic Myelogenous Leukemia (CML), the quantitation of 5 protein spots identified as beta- and/or gamma-Actin proteins, to differentiate between patients who will respond to or resist treatment when the patients are subsequently treated with an Ab1 kinase inhibitor.

The invention discloses a method which is used for extracting and separating whole protein of rat hippocampus and is suitable for dimensional electrophoresis. The method comprises the following steps: (1) placing a rat in frozen artificial cerebrospinal fluid; (2) pouring liquid nitrogen in a mortar for precooling before grinding; (3) standing turbid liquid at room temperature; (4) subpackaging liquid supernatant; (5) adding RB ( rehydration buffer) into protein conglomeration; (6) making a standard curve by 5, 10, 15, 20, 25, 30 and 35 micrograms of BSA (Bull Serum Albumin); (7) loading analysis gel, namely silver staining, with the quantity shown in the specification; (8) taking out a dried gel strip and rewarming at room temperature; (9) adding coverage oil into an isoelectric focusing disk; (10) performing isoelectric focusing treatment; (11) rewarming the treated gel strip; (12) preparing low melting agarose from upper tank liquid; and (13) carrying out coomassie blue staining on the gel and drawing. The method is easy to implement and is suitable for extracting of rat hippocampus tissue protein samples, the operation is simple and convenient, and a dimensional electrophoretogram with more protein spots, clear horizontal stripes and good repeatability is obtained.

The invention discloses a method used for measuring influences of long term applications of high and low nitrogen fertilizers on corn kernel albumin. According to the method, proteomic method is adopted to study long term positioned fertilization of high and low nitrogen fertilizers on summer corn Luyu 16 kernel albumin. It is shown by results that: the albumin content of a reference group is higher than those of two processed groups, and obvious difference strips are observed in quantificative unidirectional SDS-PAGE. It is shown by dimensional electrophoresis results that when a long term low nitrogen treated group is compared with the reference group, 7 differential protein spots are detected, one up-regulation is detected, and 6 down-regulation is detected; when a long term high nitrogen treated group is compared with the reference group, 6 differential protein spots are detected, one up-regulation is detected, and 5 down-regulation is detected. Mass spectrum identification and function classification are adopted, it is found that the proteins are a protein and amino acid metabolism associated protein, an energy metabolism associated protein, an expression regulation and control and defense and stress resistance associated protein, a cell metabolism associated protein, and a protein with unknown functions.

The invention discloses a protein extraction and electrophoresis method facilitating efficient analysis of a protein map on expression quantity differential protein, and belongs to the field of protein map engineering. Protein of aspergillus niger can be efficiently extracted and protein spots can be fully separated with the protection extraction and electrophoresis method. Aspergillus niger is broken by low-temperature wall-breaking liquid for protein extraction, and the formula of the low-temperature wall-breaking liquid is as follows: 20g of absolute ethyl alcohol, 5g of trichloroacetic acid, 5g of acetic acid, 0.02g of cetylpyridinium, 0.03g of lauryl sodium sulfate and distilled water with a constant volume of 1L. The wall-breaking liquid can increase the extraction ratio of protein. The electrophoresis method is three-dimensional electrophoresis which contributes to protein extraction of aspergillus niger and differentiation of protein expressions.

The invention relates to marker membrane protein for purifying and enriching intradermal multipotentiality cells. The protein is obtained by adopting the following steps of: in vitro culturing and amplifying cells from dermis, and diluting and culturing by using a 96 orifice plate to obtain a plurality of monoclonal cells; carrying out in vitro induction of forming fat, forming bones and forming nerves on the monoclonal cells and identifying the differentiation capacity of the cells; respectively separating and purifying two-way cloned cells and three-way cloned cells to obtain corresponding membrane protein components; finding out difference protein spots of three-way clone and two-way clone through a two-way electrophoretic analysis; carrying out LTQ mass spectrum analysis to obtain matched protein; and finding out the marker membrane protein through Western-Blot detection. The invention also relates to an application of the marker membrane protein. The marker membrane protein which is primarily screened by adopting the method and presents more obvious up-regulation or down-regulation along with the change of the differentiation capacity can be used for separating and purifying cells with multipotentiality.

The invention belongs to the technical field of proteomic technology, and particularly discloses an extracting method for phosphorylated protein suitable for two-dimensional electrophoresis of plasmamembranes of rice leaves for the first time. The extracting method comprises the steps of extraction for coarse plasma membranes of the rice leaves, purification of the plasma membranes of the rice leaves, enrichment of the phosphorylated protein of the plasma membranes of the rice leaves and the like. The extracting method is simple in operation, high in extracting efficiency and low in cost, thepurity of the plasma membranes of the rice leaves can reach 93% or above, and the yield of the enriched phosphorylated protein of the plasma membranes from protein of the plasma membranes is 3.33-3.89%. The extracting method is suitable for the plant materials with difficultly extracted phosphorylated protein of the plasma membranes and fewer interferents; the obtained phosphorylated protein of the plasma membranes contains fewer impurities and interferents and can meet the requirements of two-dimensional electrophoresis, the obtained two-dimensional electrophoretogram is clear in background,uniform in protein spot distribution and fewer in longitudinal trailing and horizontal transverse grain phenomenon and has good repeatability, and the extracting method is suitable for preparation and analysis of phosphorylated proteomes of the plasma membranes of monocotyledons.

The invention relates to a method of extracting a bovine mycoplasma outer membrane protein suitable for proteomics. The method comprises the following steps: culturing and collecting a strain, grinding by liquid nitrogen, carrying out constant-temperature ultrasonic concussion, centrifugally collecting protein precipitate, rinsing with ethanol and acetone, then carrying out centrifugal drying by virtue of vacuum freezing to obtain protein powder, and carrying out cracking treatment; and uniformly mixing the cracked solution, carrying out centrifugation, and maintaining supernatant to obtain the bovine mycoplasma outer membrane protein. According to the method disclosed by the invention, the protein extraction steps and used reagents are optimized, so that the extraction efficiency and purity degree of the protein are greatly improved, and impurities interfering a first dimension and a second dimension in dimensional electrophoresis are effectively removed; compared with the traditional method, the method is simple and convenient, is capable of saving labor and time, is high in protein extraction efficiency and less in disruptors and can be used for obtaining a map with high resolution, good repeatability, uniform protein spot distribution and clear protein spots, and has important significance for research on contagious bovine pleuropneumonia mycoplasma omics and screening research of immune proteins.

The invention provides a method for determining the type of a microtubule-associated protein-1 light chain 3 protein spot and belongs to the technical field of cell imaging. The method comprises the following steps of: observing whether a complete and rapid exchange behavior exists between the microtubule-associated protein-1 light chain 3 protein (FP-LC3) spot link-coupled with a fluorescent protein and FP-LC3 in cell plasma of a mammal; if so, determining that the FP-LC3 spot is in the type of a protein aggregate; and if few or no exchange behavior exists between the FP-LC3 and the FP-LC3 in the cell plasma of the mammal, determining that the FP-LC3 spot is in the type of an autophagy structure. By using the method, the type of the LC3 can be accurately and rapidly determined.

A detection method for skin mucus functional protein of marine fish in high temperature environment comprises the following steps: firstly collecting skin mucus of fish, then crudely extracting skin mucus protein; obtaining a skin mucus protein pattern by two-dimensional electrophoresis technology; performing image analysis by an image analysis software of Imagemaster 2D Platinum 6.0 to locate a protein spot with significant difference; performing MALDI-TOF-TOF mass spectrometry, identifying the protein spot by PMF technology and retrieval tools provided by MASCOT, NCBI websites so as to obtain functional protein related to high temperature stress. The invention uses a simple and aseptic plastic pipette to collect mucus, and the method is quick, simple, and is applicable to quick split charging; the mucus can be preserved for more than one year at a temperature below -80 DEG C; The two-dimensional electrophoresis isoelectric focusing electrophoresis procedure in the invention can completely remove salt ions, and the obtained protein spot is clear and smooth, and can be used to locate and quantify the skin mucus functional protein more accurately. The invention is applicable to detection technology such as the differential expression and the functional protein determination of skin mucus protein of marine fish in high temperature environment.