1017 results about "Polyacrylamide gel electrophoresis" patented technology

Specificity in immune responses is in part controlled by the selective interaction of T cell receptors with their cognate ligands, peptide / MHC molecules. The discriminating nature of this interaction makes these molecules, in soluble form, good candidates for selectively regulating immune responses. Attempts to exploit soluble analogs of these proteins has been hampered by the intrinsic low avidity of these molecules for their ligands. To increase the avidity of soluble analogs for their cognates to biologically relevant levels, divalent peptide / MHC complexes or T cell receptors (superdimers) were constructed. Using a recombinant DNA strategy, DNA encoding either the MHC class II / peptide or TCR heterodimers was ligated to DNA coding for murine Ig heavy and light chains. These constructs were subsequently expressed in a baculovirus expression system. Enzyme-linked immunosorbant assays (ELISA) specific for the Ig and polymorphic determinants of either the TCR or MHC fraction of the molecule indicated that infected insect cells secreted approximately 1 .mu.g / ml of soluble, conformnationally intact chimeric superdimers. SDS PAGE gel analysis of purified protein showed that expected molecular weight species. The results of flow cytometry demonstrated that the TCR and class II chimeras bound specifically with high avidity to cells bearing their cognate receptors. These superdimers will be useful for studying TCR / MHC interactions, lymphocyte tracking, identifying new antigens, and have possible uses as specific regulators of immune responses.

A crystallizable, purified chondroitinase ABC having a molecular weight of about 100,000 dalton by the measurement of the SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and the measurement by the gel permeation chromatography method, having alanine as the N-terminal amino acid and proline as the C-terminal amino acid. A process for the purification of the crystallizable purified chondroitinase ABC comprising removing nucleic acid from an surfactant solution extract obtained from cells of chondroitinase ABC-producing microorganisms and chromatographically treating by concentration gradient elution using a weak cation exchange resin or a strong cation exchange resin. A composition comprising a chondroitinase and serum albumin, gelatin, or a nonionic surfactant.

In this invention, a biomarker discovery method has been developed using specific biotin-labeled oligonucleotide ligands and magnetic streptavidin beads. In one embodiment, the oligonucleotide ligands are firstly generated by whole-cell based SELEX technique. Such ligands can recognize target cells with high affinity and specificity and can distinguish cells that are closely related to target cells even in patient samples. The targets of these oligonucleotide ligands are significant biomarkers for certain cells. These important biomarkers can be captured by forming complexes with biotin-labeled oligonucleotide ligands and collecting the complexes using magnetic streptavidin beads, whereupon the captured biomarkers are analyzed to identify the biomarkers. Analysis of biomarkers include HPLC-Mass Spectroscopy analysis, polyacrylamide gel electrophoresis, flow cytometry, and the like. The identified biomarkers can be used for pathological diagnosis and therapeutic applications. Using the disclosed methods, highly specific biomarkers of any kinds of cells, in particular cancer cells, can easily be identified without prior knowledge of the existence of such biomarkers.

The invention relates to a method for separating and purifying mussel mucin by using a mixed adsorption chromatography. Mussel mucin contains a group of L-3,4- -Dihydroxyphenylalanine (L-DOPA), a phenohydroxyl group thereof can act as the supplier for hydrogen bond, the benzene ring thereof can generate a hydrophobic effect, and the lysine thereof with strong positive charges is capable of forming a static bond. On the basis of the properties of mussel mucin, a mixed adsorption chromatography (i.e. the adsorption chromatography based on three principles of adsorption with hydrogen bond, adsorption with hydrophobic effect, and static adsorption) is adopted to overcome the problem of low yielding rate of mussel mucin in the prior art for separating and purifying mussel mucin. An strong acid extraction is adopted to eliminate small-molecular compounds from a desalting column, an argar medium with high concentration and high cross-linking degree to separate and purify mussel mucin, and an acetic acid-urea- polyacrylamide gel electrophoresis is used to differentiate mussel mucin through specific chromogenesis with nitro blue tetrazolium. Three principles adopted with one separation medium to separate mussel mucin achieve high selectivity, simplify the purification technology, and decrease production cost.

The invention relates to a method for separating and purifying the sea-mussel mucin by using salting out and dialyzing, the salting out and the dialyzing utilizes the proteinaceous physicochemical character to purify by a non- chromatograph method, and solves the problem of high apparatus and material cost in the chromatograph method for purifying the sea-mussel mucin, a quick, low-cost and high-concentration method for separating and purifying the sea-mussel mucin is provided. The strong acid is used for extracting, the salting out and the dialyzing are used for purifying, the sea-mussel mucin is determined by adopting the acetic acid-carbamide- polyacrylamide gel electrophoresis via the blue tetrazolium specificity colour development, and the purity is determined by the inversed phase chromatography quantitatively.

A protein which inhibits osteoclast diffraction and / or maturation and a method for producing the protein. The protein is produced by human embryonic lung fibroblasts and has a molecular weight of about 60 kD and about 120 kD under non-reducing conditions and about 60 kD under reducing conditions an SDS-polyacrylamide gel electrophoresis. The protein can be isolated and purified from the culture medium of fibroblasts. Furthermore, the protein can be produced by gene engineering. The present invention includes cDNA for producing the protein by gene engineering, antibodies having specific affinity for the protein or a method for determining protein concentration using these antibodies.

Plant cell expansion is regulated by wall relaxation and yielding, which is thought to be catalyzed by elusive "wall loosening" enzymes. By employing a reconstitution approach, we initially found that a crude protein extract from the cell walls of growing cucumber seedlings possessed the ability to induce the extension of isolated cell walls. This activity was restricted to the growing region of the stem and could induce the extension of isolated cell walls from various dicots and monocots, but was less effective on grass coleoptile walls. Sequential HPLC fractionation of the active wall extract revealed two proteins with molecular masses of 29 and 30 kD, as measured by SDS-PAGE, associated with such activity. Each protein, by itself, could induce wall extension without detectable hydrolytic breakdown of the wall. We proposed the name "expansins" for this class of proteins. Expansins have been isolated from various plant sources including oat, cucumber, broccoli, celery, tomato, cotton, cabbage, and corn, and also from snail and its feces. These proteins weaken the intermolecular bonds between plant wall polysaccharides. They decrease the mechanical strength of commercial products made from polysaccharides, such as paper, and therefore present a novel approach in developing new technologies in industries which make use of such polysaccharides, such as in the paper industry, in the applications of polysaccharide gums and related products. These proteins moreover present a novel approach in the control of plant growth.

The present invention is concerned with non-soluble proteins and soluble or insoluble fragments thereof, which bind TNF, in homogeneous form, as well as their physiologically compatible salts, especially those proteins having a molecular weight of about 55 or 75 kD (non-reducing SDS-PAGE conditions), a process for the isolation of such proteins, antibodies against such proteins, DNA sequences which code for non-soluble proteins and soluble or non-soluble fragments thereof, which bind TNF, as well as those which code for proteins comprising partly of a soluble fragment, which binds TNF, and partly of all domains except the first of the constant region of the heavy chain of human immunoglobulins and the recombinant proteins coded thereby as well as a process for their manufacture using transformed pro- and eukaryotic host cells.

A protein from M. catarrhalis, designated the 74 kD protein, is isolated and purified. The 74 kD protein has an amino-terminal amino acid sequence which is conserved among various strains of M. catarrhalis. The protein has a molecular weight of approximately 74.9 kD as measured on a 10% SDS-PAGE gel, while its molecular weight as measured by mass spectrometry is approximately 74 kD. The 74 kD protein is used to prepare a vaccine composition which elicits a protective immune response in a mammalian host, to protect the host against disease caused by M. catarrhalis.

The invention relates to a method for identifying the animal-based components in meat and meat products by utilizing an SDS-PAGE (Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis) method, and belongs to the field of food sanitation inspection. The method provided by the invention is utilized to identify the pig, cattle, sheep, chicken and fish-based components in the meat products; the SDS-PAGE is carried out on the soluble proteins which are extracted from the processed sample by carrying out different temperature thermal treatments on fresh animal muscle tissues; and the pig, cattle, sheep, chicken and fish-based components in the meat products are judged according to an electrophoresis pattern. The method provided by the invention has the characteristics of strong specificity and sensitivity, and visual and reliable result judgment, and is simple to operate and has the extremely important significances on preventing animal disease pathogens such as foot-and-mouth disease, avian influenza and the like from spreading into China, preventing illegal retailers from concealing commodity components and guaranteeing the health of people.

The invention provides diacylglycerol acyltransferase (DAGAT) proteins, wherein said proteins are active in the formation of triacylglycerol from fatty acyl and diacylglycerol substrates. In one aspect, Mortierella ramanniana DAGAT proteins have been isolated and have molecular weights of between approximately 36 and 37 kDa as measured by SDS-PAGE. The invention also provides novel DAGAT polynucleotide and polypeptide sequences and to methods of producing such polypeptides using recombinannt techniques. In addition, methods are provided for using such sequences to alter triacylglycerol levels in plants and to treat diseases associated with altered DAGAT activity or expression.

The present disclosure relates to a method for metabolic oligosaccharide engineering that incorporates derivatized alkyne-bearing sugar analogs as “tags” into cellular glycoconjugates. The disclosed method incorporates alkynyl derivatized Fuc and alkynyl derivatized ManNAc sugars into a cellular glycoconjugate. A chemical probe comprising an azide group and a visual probe or a fluorogenic probe is used to label the alkyne-derivatized sugar-tagged glycoconjugate. In one aspect, the chemical probe binds covalently to the alkynyl group by Cu(I)-catalyzed [3+2] azide-alkyne cycloaddition and is visualized at the cell surface, intracellularly, or in a cellular extract. The labeled glycoconjugate is capable of detection by flow cytometry, SDS-PAGE, Western blot, ELISA or confocal microscopy, and mass spectrometry.

There are provided a soluble CD14 antigen which is a novel in vivo protein useful as a marker for diagnosing sepsis and has the following characteristic features 1) to 3):1) a molecular weight of 13±2 kDa when measured by SDS-PAGE under non-reducing conditions;2) an amino acid sequence in which the amino acid sequence of SEQ ID NO:1 is present on its N terminal; and3) ability to specifically bind to an antibody prepared by using a peptide comprising 16 amino acid residues described in SEQ ID NO:2 for the antigen; and a recombinant soluble CD14 fragment.

There are provided natural rubber free from substances which cause Type I allergy, a rubber compositions having good processability and physical properties which comprises the natural rubber and other rubber, and a tire product comprising the natural rubber. The natural rubber of the present invention exhibits the above properties by containing substantially no proteins specified by the bands of 14, 31 and 45 kDa by SDS-PAGE.

The invention discloses an anti-rabies virus monoclonal antibody and a preparation method and an application, belonging to the field of biomedicine and particularly relating to the preparation of a monoclonal antibody capable of identifying rabies virus and the application. The monoclonal antibody of the invention is screened by indirect Enzyme-linked immunosorbent assay (ELISA), and specificity and affinity thereof combined with antigen are identified by methods such as polyacrylamide gel electrophoresis analysis, speckle ELISA, immunoblot analysis and the like. The anti-rabies virus monoclonal antibody of the invention can be applied in multiple testing methods of antigen of rabies virus and can be also applied in the preparation of rabies virus detecting kit. The anti-rabies virus monoclonal antibody is secreted by anti-rabies virus monoclonal antibody hybridoma cell strain 2C5 with the preservation number being CGMCC No.3014.

The present disclosure relates to a method for metabolic oligosaccharide engineering that incorporates derivatized alkyne-bearing sugar analogs as “tags” into cellular glycoconjugates. The disclosed method incorporates alkynyl derivatized Fuc and alkynyl derivatized ManNAc sugars into a cellular glycoconjugate. A chemical probe comprising an azide group and a visual probe or a fluorogenic probe is used to label the alkyne-derivatized sugar-tagged glycoconjugate. In one aspect, the chemical probe binds covalently to the alkynyl group by Cu(I)-catalyzed [3+2] azide-alkyne cycloaddition and is visualized at the cell surface, intracellularly, or in a cellular extract. The labeled glycoconjugate is capable of detection by flow cytometry, SDS-PAGE, Western blot, ELISA or confocal microscopy, and mass spectrometry.

A water soluble extract of M.charantia named MC6, methods for its preparation and methods for its use in the treatment of hyperglycemic disorders are provided. The active MC6 is characterized by moving as a single band on SDS-PAGE having a molecular weight of less than 10 kDal, and by comprising three peptides. Also provided is a peptide component of MC6 named MC6.1, as well as analogues and mimetics thereof. The active MC6, MC 6.1, MC6.2, and MC6.3 exhibit hypoglycemic activity, even following oral administration. Also provided are methods of using the active agents to treat hyperglycemic disorders, particularly diabetes, where the active agents are preferably orally administered.

This invention is intended to isolate and identify a vWF-specific cleaving protease.The vWF-specific cleaving protease cleaves a bond between residues Tyr 842 and Met 843 of vWF and comprises a polypeptide chain having Leu-Leu-Val-Ala-Val as a partial sequence, and more preferably comprises a polypeptide chain having the partial N-terminal amino acid sequence of a mature protein, Ala-Ala-Gly-Gly-Ile-Leu-His-Leu-Glu-Leu-Leu-Val-Ala-Val, and having a molecular weight of 105 to 160 kDa in SDS-PAGE under reducing or non-reducing conditions. Isolation and identification of this vWF-specific cleaving protease have led to the possibility of replacement therapy for patients having diseases resulting from a deficiency of the protease, such as thrombotic thrombocytopenic purpura.

The present invention relates to a method for detecting immunoblotting based on quantum dot labeling. Firstly, connecting quantum dot with carboxyl or amido with second antibody by chemical reaction to form compound with quantum dot label; employing polyacrylamide gel electrophoresis, analytes is protein, probe is antibody, quantum dot labeled second antibody is used to show color. Protein sample separated by PAGE is transferred to solid carrier, the protein or polypeptide on solid carrier as antigen is processed immunoreaction with corresponding antibody, and then reacted with quantum dot labeled second antibody, under the UV lamp, quantum dot emitting fluorescence to detect electrophoresis separated specific objective gene expression protein component. Immunoblotting image is gained by filter equipped of simple UV imaging system. The present invention employs quantum dot connected with second antibody as excellent fluorescence imaging agent of immunoblotting to enhance the sensitivity and precision.

The invention discloses a recombinant antigenic protein for diagnosing echinococcosis granulosus (having an amino acid sequence represented by SEQIDNo.1). In addition, the invention also discloses the preparation method of the recombinant antigenic protein, which comprises: amplifying an EgEPC1 gene by using RT-PCR; cloning the EgEPC1 gene in a pGEM-T vector; connecting the EgEPC1 gene with an expression vector PET28a(+) to form a recombinant plasmid PET28a-EgEPC1; transforming the recombinant plasmid PET28a-EgEPC1 to Escherichia coli BL21(DE3) and expressing the recombinant protein through IPTG induction; and identifying a purified recombinant antigen by using SDS-PAGE and Western blotting. In addition, the invention also discloses the diagnosis use of the recombinant antigenic protein. Experiments show that the recombinant antigenic protein of the invention has the advantages of high sensibility and specificity for the diagnosis of echinococcosis granulosus, and has a promising application prospect in the diagnosis of the echinococcosis granulosus.

The invention provides a construction method for a DNA fingerprint database of a high flux cotton variety. The method comprises the following steps: extracting DNAs of the cotton variety, carrying out SSR-PCR amplification by using 40 pairs of primers (with nucleotide sequences as represented by SEQ ID No. 1-80), carrying out fluorescence multiple PCR combination on products obtained after amplification, carrying out capillary five-color fluorescence electrophoresis detection with a DNA analyzer and constructing the DNA fingerprint database of the cotton variety according to a fluorescence spectrum. Compared to detection efficiency of conventional polyacrylamide gel electrophoresis, detection efficiency of the method provided in the invention is increased by more than 10 times; in addition, since the size of fragments of the amplification products is obtained through automatic comparison of internal standard of molecular weight, human errors in plate reading is reduced, and accuracy and reliability of data results are utmostly guaranteed at the same time.

A protein which inhibits osteoclast differentiation and / or maturation and a method of production of the protein. The protein is produced by human embryonic lung fibroblasts and has molecular weight of about 60 kD and about 120 kD under non-reducing conditions and about 60 kD under reducing conditions on SDS-polyacrylamide gel electrophoresis, respectively. The protein can be isolated and purified from culture medium of the said fibroblasts. Furthermore, the protein can be produced by gene engineering. The present invention includes cDNA for producing the protein by gene engineering, antibodies having specific affinity to the protein or a method for determination of the protein concentration using the antibodies.

The invention provides a method for establishing an SSR (Simple Sequence Repeat) fingerprint spectrum of broad beans, and belongs to the technical field of plant variety identification and breeding. The method provided by the invention provides a marker combination consisting of 21 pairs of broad bean genome SSR primers, and the nucleotide sequences of which are shown in SEQ ID NO. 1 to 42 respectively; the broad bean varieties are identified by PCR (Polymerase Chain Reaction) amplification technology, non-denaturing polyacrylamide gel electrophoresis or capillary electrophoresis detection technology, and the identification of broad bean seed varieties and genetic diversity assessment work can be completed in a short time. The method has the advantages of time saving, high efficiency, fastness, accuracy, and convenience in operation, and the identification results are not easily affected by the environment. The method provided by the invention has the advantages of effectively monitoring the authenticity of the broad bean seeds, revealing the genetic variation and genetic relationship of the varieties from the DNA level, protecting the crop varieties, preventing the fake and inferior varieties from entering the market, and also providing a technical support for the reasonable utilization of excellent germplasm during the breeding of broad bean varieties, and has good application prospects.

The present invention discloses a cabbage SSR fingerprint construction method, and belongs to the technical field of agricultural cabbage breeding and application. DNA of a variety to be tested is extracted. Twenty nine pairs of core SSR primers as shown in a sequence table SEQ ID No. 1-58 are respectively used for PCR amplification of the DNA of the variety to be tested. PCR products are detected by a denatured polyacrylamide gel electrophoresis or a five-color fluorescent capillary electrophoresis to construct cabbage variety SSR fingerprints. The invention establishes a fast and accurate, economic and simple, stable and reliable cabbage DNA fingerprint certification standard experimental system, and on this basis, develops a DNA fingerprint detection method applicable for cabbage variety identification. The present invention utilizes capillary electrophoresis methods for variety identification of cabbage, enhances the experimental efficiency in time, and has the advantages of fast speed, accuracy and easy operation, and identification results are not influenced by environments.

The present invention uses an optimized albumen extracting method, can test the constituent and the content of the foam albumen in the beer barley and the malt using the dodecyl sodium sulfate - polyacrylamide gel (SDS-PAGE) electrophoresis technology, and can detect the change of the constituents and the content of the foam albumen during the germinating process of the beer barley and the malt.

The invention relates to a glycosylated modifying method for improving the antioxidant activity of a whey protein. The method comprises the following step of: by using xylose, glucose, fructose, lactose, maltose, saccharose and the whey protein as reaction raw materials, performing a damp and hot glycosylated reaction according to a certain proportion. The result shows that the reducing capacity of a whey protein-xylose compound, the DPPH free radical scavenging capacity, the.OH free radical scavenging capacity and the anti-lipid peroxidation capacity are improved to the maximum extent along with increase of pH. SDS-PAGE displays that the molecular weight of the whey protein-xylose compound after reaction is remarkably increased. Fourier Transform Infrared Spectroscopy (FTIR) results show that amides I and II in the whey protein-xylose compound are remarkably reduced. The method provided by the invention aims to obtain the glycosylated modifying method for the whey protein with stronger antioxidant function. The antioxidant function characteristic of the whey protein is improved, a novel functional whey protein ingredient is developed, and the application range is expanded and a novel functional additive is increased.

The invention discloses a method for constructing a wheat SSR fingerprint, which belongs to the field of agricultural wheat breeding and application technologies. The method comprises the following steps: extracting DNA (deoxyribonucleic acid) of a wheat variety to be tested; carrying out PCR (polymerase chain reaction) amplification to the DNA Of the wheat variety to be tested by using 13 pairs of basic core SSR fluorescent dye primers with the ID No.1 to 26 and shown in a sequence table SEQ and 12 pairs of extended core SSR fluorescent dye primers with the ID No.27 to 50 and shown in the sequence table SEQ respectively; and detecting PCR amplification products by denaturing polyacrylamide gel electrophoresis or five-color fluorescent capillary electrophoresis, and constructing the SSR fingerprint of the wheat variety. The method provided by the invention can identify the wheat variety in a laboratory by using technologies of the PCR amplification, the gel electrophoresis and capillary electrophoresis according to the polymorphism of SSR sites, can greatly improve the test efficiency in time, and has the advantages of high speed, high accuracy, convenience for operation and the like; and the identification result is not liable to be affected by the environment.

Bacillus sp. NTAP-1 having been deposited under accession number FERM BP-6926; and a collagen-decomposing enzyme produced by bacterium. The above enzyme (1) has a capability of hydrolyzing, at the highest efficiency, collagen and gelatin from among casein, gelatin, albumin and collagen; (2) shows the optimum pH of 3.5 to 4.5; (3) shows the optimum temperature of 65 to 70° C.; (4) after heating at 60° C. at pH 6.0 for 4 hours, sustains an activity amounting to 60% or more of the level before the heat treatment; (5) remains stable at pH 3 to 6; and a molecular weight of approximately 46,000 when measured by SDS-PAGE.

The invention discloses a recombinant antigen protein (of which the amino acid sequence is shown as SEQ ID NO:1) for diagnosing echinococcosis granulose. Moreover, the invention further discloses a preparation method of the recombinant antigen protein. The method comprises the following steps of: amplifying an EgENO gene by adopting RT-PCR (Reverse Transcription-Polymerase Chain Reaction); cloning the EgENO gene into an expression vector PET28a (+) for constructing a recombined plasmid PET28a-EgENO; converting into escherichia coli BL21(DE3); inducing the expression of a recombinant protein through IPTG (Isopropyl-beta-d-Thiogalactoside); and identifying a purified recombinant antigen by using SDS-PAGE (Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis) and Western blotting. In addition, the invention further discloses a diagnosis application of the recombinant antigen protein. As proved by an experiment, the recombinant antigen protein has the advantages of high sensitivity, high specificity and the like for the diagnosis of the echinococcosis granulosa, and has a wide application prospect on the aspect of diagnosis of the echinococcosis granulosa.