71 results about "GTP-sepharose" patented technology

The present invention relates to a method and apparatus for forming agarose or cored agarose beads. The process involves dissolving / gelation the agarose in a suitable liquid, mixing it with a hydrophobic liquid to form an emulsion and maintaining that emulsion at a temperature equal to or greater than the gelation point of the agarose, passing it through a static mixer to create agarose droplets and solidifying the agarose droplets in a second bath of hydrophobic liquid. The beads can then be washed and used or further processed to crosslink the agarose and / or add various functionalities on to the agarose. Another method for solidifying the agarose droplets is by using a heat exchanger to cool the stream continuously after it exits the static mixer. A similar process is used for the “cored” beads except cores, preferably in bead form, are introduced to the agarose before it enters the first hydrophobic liquid so that the agarose forms a coating on the cores. A similar process with either agarose beads (made by this or another process) or cored agarose (made by this or another process) can be used to add multiple layers of agarose on to the existing beads. An apparatus for running the process is also disclosed.

The present invention provides processes for the production of preformed albumin conjugates. In particular, the invention provides processes for the in-vitro conjugation of a therapeutic compound to recombinant albumin, wherein a therapeutic compound comprising a reactive group is contacted to recombinant albumin in solution to form a conjugate. The processes provide for conjugation to albumin species of increasing homogeneity . The resulting conjugate is purified by chromatography, in particular hydrophobic interaction chromatography comprising phenyl sepharose and butyl sepharose chromatography.

The invention discloses a preparation method of magnetic sepharose gel microspheres. The preparation method comprises the following steps: (1) preparing ferrous powder with scattered active agent: adding the ferrous powder into surface active agent water solution, heating and mechanically stirring; washing and drying the obtained product to obtain the ferrous powder with scattered active agent; and (2) preparing the magnetic sepharose gel microspheres: dissolving an emulsifying agent span80 into liquid paraffin for preheating; taking sepharose gel and the ferrous powder with scattered active agent, adding secondary distilled water to prepare sepharose solution, conducting ultrasonic scattering and heating and dissolving; adding the dissolved sepharose solution into span80-containing liquid paraffin solution; after the reaction is finished, cooling a sample and conducting slow and mechanical stirring; separating the sample, and washing to obtain the magnetic sepharose gel microspheres. By adopting a reversed phase suspension embedding method, the prepared magnetic sepharose gel microspheres are strong in magnetic responsibility, and have a good effect on the separation and purification of protein.

The invention discloses a method for detecting the cytotoxicity of a to-be-tested medicine to a target cell and a cell chip specially used by the method. The cell chip provided by the invention comprises a supporting medium and a sepharose gel film attached to the upper part of the supporting medium, wherein a plurality of microtraps are distributed in the sepharose gel film, and the size of each microtrap is matched with that of the individual target cell; and the individual target cell is attached to the inside of each microtrap. The experiment proves that the cell chip provided by the invention can detect the cytotoxicity of the to-be-tested medicine to the cell and / or detect the gene damage of the to-be-tested medicine to the cell and / or detect the influences of the to-be-tested medicine to the expression of interest protein in the cell and / or screen high-content drugs having effects for the cells. The cell chip provided by the invention has important application values.

The invention relates to a DEAE dextran-modified agarose gel-based chromatography medium and a preparation method and application thereof. The method includes the steps that first, the molecular weight of DEAE Dextran for grafting is selected; next, the concentration of a DEAE Dextran aqueous solution is controlled; then, the time of diffusion before DEAE Dextran coupling is determined; finally, the reaction time of coupled DEAE Dextran and the concentration and volume of sodium hydroxide are determined. The DEAE dextran-modified agarose gel-based chromatography medium has a strong adsorption property for protein under different modification densities, shows a high adsorption capacity and a high adsorption rate and improves separation efficiency. The medium is convenient to clean and degerm, easy to regenerate and good in biocompatibility. The preparation method is simple and harmfulless and has broad application prospects in efficient and rapid separation and purification of protein.

The invention relates to the field of gene engineering, and discloses a construction and an expression for two engineering bacteria of two human glucagon-like peptide-1 analogues, and a preparation method for the objective peptides. The objective peptides are formed by inserting objective sequence after double digestions, adding His labels at the front ends of fusion proteins, obtaining objective peptides by cutting the fusion proteins with acid via gaining the fusion proteins, and after an isoelectric precipitation, obtaining the objective peptides again through an affinity chromatography of a Ni agarose gel after an isoelectric precipitation. Freeze-dried objective peptides are dissolved in a PBS buffer solution, oral glucose tolerance test and determination of serum insulin concentration are carried out on normal ICR mice. The two analogues are modified peptides of human glucagon-like peptide-1. Experiments demonstrate that both the two peptides have effects for reducing blood glucose and can be used for treating type 2 diabetes mellitus. Furthermore, the invention also provides the preparation method for the peptides.

The invention discloses a molecular identification method of four major Chinese carps in fresh water. The method provided by the invention comprises the following steps: (1) genome DNA extraction; (2) PCR amplification; (3) PCR reaction: the reaction procedure is started with predenaturation at 94 DEG C for 5min, initial denaturation lasts for 5min, then denaturation is carried out at 94 DEG C for 30s, annealing is carried out at 62 DEG C for 30s, extension is carried out at 72 DEG C for 1min, 30 cycles are performed, and after the cycles, extension is carried out again at 72 DEG C for 8min; (4) electrophoresis detection: after PCR, a solution of 2-5 microliters is put on 1% of sepharose gel for electrophoresis, and observation and photographing are carried out; and (5) result judgment. The method provided by the invention has advantages of simple operation, accurate judgment and low cost.

The invention relates to a bilateral molecular marker identification technology of a melon wilt resistant variety. By the utilization of polymerase chain reaction and bilateral molecular markers ML430 and MR296 which are closely linked to a melon wilt resistance gene, specific DNA fragments are amplified. The size differences of amplified DNA fragments between a resistant variety and a susceptible variety can be displayed on different locations of a sepharose gel by electrophoretic technique. And the sizes and locations are compared with that of DNA fragments amplified from a standard resistant variety and a standard susceptible variety. Two varieties which have the same size or lie in the same location belong to the variety of the same type. When banding patterns of the two molecular markers both are resistance type, the variety is the resistant variety. The method provided by the invention has advantages of convenient sampling, precise identification, and no influence by plant growth and development and weather conditions. The technology provided by the invention is mainly used for the screening and identification of wilt resistance variety resources, and can also be used for molecular marker assisted breeding of melon.

The invention discloses a multi-purpose separation medium and a preparation method thereof, wherein, tetrazole is the functional group of the medium. More particularly, a separation medium with novel structure is prepared by linking the tetrazole on the surface of silica gel, dextran and agarose gel separation group. The invention has dual purposes of being applied to ion-exchange chromatography and metal chelate chromatography. When the medium is applied to the ion-exchange separation of proteins, the separation of proteins is highly selective, easy to regenerate and quick to separate and both the quality and recovery rate of the activity of the proteins are high. When the medium is applied to the metal chelate chromatography of proteins, the separation of proteins is highly selective, small in metal ion loss and quick to separate and both the quality and recovery rate of activity of the proteins are high. The invention can be applied to the quick separation and purification of genetic engineering products and plasma proteins.

The invention discloses a preparation method of an ulva fasciata delile sulfated polysaccharide with anti-tumor activity. The preparation method comprises the following steps of crushing and ultrasonically extracting ulva fasciata delile by water to prepare a crude polysaccharide through alcohol precipitation; carrying out depigmentation deproteinization by using a radial-flow chromatographic column which is filled with weakly basic anion exchange resin; carrying out ultrafiltration separation according to molecular weight after elution; and taking and adding an ulva fasciata delile polysaccharide compound with molecular weight of 10 KD to 30 KD into the radial-flow chromatographic column which is filled with DEAE (Diethyl Aminoethanol) sepharose gel for enriching and purifying, wherein the prepared polysaccharide with average molecular weight of 20 KD has remarkable antitumor activity.

The invention relates to a separating and purifying method for preparing white muscardine silkworm anticancer-activity polysaccharide BBPW-1, which comprises the following steps: degreasing white muscardine silkworm powder under reflux of acetone-petroleum ether and 80% ethanol to remove glycosides and alkaloids, and extracting with distilled water at 80-100 DEG C to obtain crude polysaccharide; and after removing proteins from the crude polysaccharide by a Sevag process, separating out the neutral polysaccharide component by DEAE (diethylaminoethanol) sepharose ion-exchange chromatography, carrying out propylene dextrangel S-300 chromatography, taking the first eluting peak, concentrating under reduced pressure, carrying out propylene dextrangel S-500 chromatography for further purification, and carrying out freeze-drying to obtain the white muscardine silkworm anticancer-activity polysaccharide BBPW-1. The product prepared by the method provided by the invention has uniform purity, and the molecular weight is 3.67*10<6>Da; and the product has an inhibiting action on growth of human cervical cancer cells Hela and human liver cancer cells HepG2, does not have any adverse effect on growth of normal human embryo kidney cells HEK293 and mouse macrophages RAW264.7, and can be used for developing anticancer products.

The invention relates to a method for chemically synthesizing oxytocin by using hydrogen-bonding adsorption chromatographic separation and purification. In the method, a high-concentration and high-crosslinking-degree agarose gel medium, on the surface of which beta-cyclodextrin ligand is bonded, is adopted; and the oxytocin is separated and purified from a rough oxytocin product which is obtained through chemical synthesis (comprising liquid-phase synthesis and solid-phase synthesis) according to a hydrogen-bonding adsorption principle. The method is used for producing oxytocin raw material medicines, has the advantages of high selectivity, high sample loading capacity, high medium recycling rate and the like; and the defects of low medium recycling rate and low sample loading capacity existing in the conventional reverse phase chromatography are overcome.

The present invention relates to a method for concentrating and / or purifying prion PrPSc proteins by contacting prion PrPSc proteins with sepharose under conditions that allow for the specific and high affinity binding of the sepharose to the prion PrPSc proteins and removing the unbound non-prion proteins from the sepharose, as well as the same method for removing prion PrPSc proteins from body fluids by contacting body fluids with sepharose under conditions that allow for the specific and high affinity binding of the sepharose to the prion PrPSc proteins and removing the body fluid from said sepharose. In addition, the present invention is directed to a method for separating and / or enriching prion PrPSc proteins from PrPC proteins by contacting prion PrPSc proteins and PrPC proteins with a ligand-modified sepharose under conditions that allow for the specific and high affinity binding of the sepharose part to the prion PrPSc proteins and the binding of the ligand part of the sepharose to PrPC proteins, adding a selective release agent to the sepharose-bound proteins under conditions that allow for the release of non-prion proteins and PrPC proteins from the ligand part of the sepharose but not for the release of the prion PrPSc proteins, and removing the non-prion proteins and PrPC from the sepharose. Another aspect of the present invention concerns the use of the before-mentioned methods for concentrating, purifying and / or removing prion PrPSc proteins.

The present invention is agarose gel-m-aminobenzene boric acid affinity chromatography for separating single glycosylated component with plasmin activity from the homogenated earthworm liquid. During the affinity chromatography, agarose gel is used as stuffing, m-aminobenzene boric acid as aglycone and cis-monosaccharide as specific eluant, and the glycosylated component is produced through dialysis or molecular sieve process to desalt, drying and storing. The single component, SDS-PAGE detection shows, has N-terminal sequence N-Ala-Glu-Val-Cys-Cys-Property-Asp-Ile- and is one new kind of protein. The chromophoric substrate activity test shows that it has relatively high plasmin activity. The said process is simple, convenient, low in cost and suitable for large-scale production.

The invention relates to a dual PCR method for detecting theileria hirci and anaplasma. The method includes the steps of finding a gene conserved seuqnece target gene locus and designing a specific primer sequence I according to the 18S rRNA gene sequence of theileria hirci, finding a gene conserved seuqnece target gene locus and designing the specific primer sequence II according to the 16S rRNA gene sequence of anaplasma, adding the upstream primer and the downstream primer of the specific primer sequence I and the upstream primer and the downstream primer of the specific primer sequence II and PCR mixed liquid into a PCR pipe, mixing the materials and adding theileria hirci DNA and anaplasma DNA extracted from a to-be-tested blood sample to the mixture, putting the PCR pipe into a PCR amplification instrument to be circulated, obtaining a PCR product, and putting the PCR product on 2% sepharose gel to be subjected to electrophoresis. The method has the advantages of being rapid, specific, sensitive, efficient, low in cost and the like and is beneficial to clinical application.

The method is used for separating nucleic acids and other similar constructs. It involves selective introduction, enhancement, or stabilization of affinity handles such as single-strandedness in the undesired (or desired) nucleic acids as compared to the usual structure (e.g., double-strandedness) of the desired (or undesired) nucleic acids. The undesired (or desired) nucleic acids are separated from the desired (or undesired) nucleic acids due to capture by methods including but not limited to immobilized metal affinity chromatography, immobilized single-stranded DNA binding (SSB) protein, and immobilized oligonucleotides. The invention is useful to: remove contaminating genomic DNA from plasmid DNA; remove genomic DNA from plasmids, BACs, and similar constructs; selectively separate oligonucleotides and similar DNA fragments from their partner strands; purification of aptamers, (deoxy)-ribozymes and other highly structured nucleic acids; Separation of restriction fragments without using agarose gels; manufacture recombinant Taq polymerase or similar products that are sensitive to host genomic DNA contamination; and other applications

The invention discloses a protein A immunoadsorbing material for absorbing antibodies, which is a polymer material coupled with agarose gel and protein A, and the protein A is a recombinant protein A with cysteine; The protein A immunosorbent material is prepared through three steps of preparation of recombinant protein A, activation of agarose gel, and synthesis of the immunosorbent material; the protein A immunosorbent material of the present invention can be directly, efficiently and selectively adsorbed from body fluids Pathogenic antibodies related to autoimmune diseases can be safely performed in extracorporeal circulation, and the purpose of eliminating pathogenic antibodies in patients can be achieved by one-time perfusion.

The invention relates to an affinity chromatography medium for separating and purifying a GABA receptor and a preparation method thereof. The technological process comprises the following steps: carrying out chemical modification on fipronil to obtain an affinity ligand; by taking agarose gel as a matrix, after activating by an epoxy activating agent, coupling the fipronil affinity ligand to obtain an affinity medium; packing the affinity medium so as to be used for separating and purifying fish GABA receptor protein. The affinity chromatography medium synthesized according to the invention has the advantages of being good in selectivity, strong in specificity, high in mechanical strength and good in separation effect; in addition, the preparation process is simple and the magnifying is easy.

The invention relates to a method for preparing a series of N-desulfated heparin derivative affinity chromatographic materials. The method comprises the following steps of: by respectively utilizing a de-2-O desulfated heparin derivative, a de-6-O desulfated heparin derivative and a de-N-desulfated and acetylated heparin derivative as ligands and using sepharose particles as a carrier, preparing a series of stable N-desulfated heparin derivative affinity chromatographic materials through the reductive amination reaction of the activated sepharose particles and all the heparin derivatives in the presence of a strong reducing agent. According to the method, heparin derivatives with different desulfated loci are used for preparing the affinity chromatographic materials, so that three heparin derivative affinity chromatographies with high stability and high activity are obtained; and the affinity chromatographic materials can be used for researching the structure and functions of heparin and separating and concentrating functional protein, tumor cells, virus and the like.

The invention provides a multi-PCR kit for rapidly identifying Dragonfish type and an identifying method thereof; the kit includes 1, three upstream primers and three downstream primers; 2, positive comparison product; 3, PCR reaction fluid; meanwhile, the method also provides a method for rapidly identifying the Dragonfish type, wherein the method includes steps of 1), selecting three upstream primers and three downstream primers; 2), preparing a gene group DNA template; 3), respectively adding the upstream primers and the downstream primers in the gene group DNA template prepared by the positive controlled DNA template and the Dragonfish to be tested, so as to perform PCR amplified reaction;4). Taking 5 miu L products after amplification and applying samples to 2.5% of sepharose gel; taking 50 bp Marker (stripes 50, 100, 150, 200, 250, 300, 350, 400, 450, 500) as standard molecule reference and performing electrophoresis; applying the electrophoresis result to the analysis of an ultraviolet gel imaging system; 5), judging result. The kit and the method can improve the identifying efficiency, save time, and save the identifying cost.

The invention relates to a method for separating and purifying polypeptide through a hydrogen binding adsorption chromatographic medium of quercitin aglucon and high-concentration and high-crosslinking degree agarose gel. The quercitin aglucon and the high-concentration and high-crosslinking degree agarose gel serve as a medium of a matrix; and the high-purity polypeptide is further separated and purified from a biological tissue extract, a protein hydrolysate or a polypeptide synthesis mixture based on the principle of the hydrogen binding adsorption chromatography characterized by hydrogen binding mutual effect. The method has the characteristics of high selectivity, simple purification process, large sample carrying amount, high medium recycling degree and the like. The defects that the traditional method has complex steps, low reversed-phase chromatographic medium recycling degree, difficult mass production, low sample carrying amount, poor biocompatibility and the like are overcome. The purity of the obtained polypeptide fractions is determined through reversed-phase high efficiency liquid chromatography, and the sequence and the molecular weight of the polypeptide are determined through a mass spectrum.

The invention relates to the technical field of gene cloning and expression, and in particular to a method for expressing and purifying a recombinant human sex hormone haptoglobin N-terminal 51-218aa.According to the invention, Escherichia coli expression and purification of the recombinant human sex hormone haptoglobin are employed to provide a construction method of a recombinant human SHBG protein N-terminal 51-218aa expression vector, and the N-terminal 51-218aa of the recombinant human SHBG protein obtain by induced expression is subjected to Ni-Sepharose chromatographic column and two-step purification. The N-terminal 51-218aa of the recombinant human SHBG protein prepared by the method of the invention has the characteristics of high yield, high purity and low cost, and is useful for further self-developing of a detection kit of the protein.

The present invention relates to the use of milk or a derivative thereof for identifying prion proteins, preferably PrPSc prion proteins, in a mammal. The present invention is also directed to a method for identifying prion proteins, preferably PrPSc prion proteins, in mammals, comprising the step of contacting milk or a derivative thereof with an agent having high affinity and selectivity for prion proteins, preferably for PrPSc prion proteins. In addition, a further aspect the present invention concerns a method for removing PrPC and / or PrPSc prion proteins, preferably PrPSc prion proteins, from milk or a milk derivative wherein milk or a derivative thereof is contacted with sepharose, preferably sepharose comprising divalent immobilized metal ions.

The invention a protein A adsorption material for targeted adsorption. A protein A is coupled to a high molecular material on a sepharose gel microspherical carrier covalently. The chemical structureof the protein A adsorption material is as follows: a formula as shown in the description represents a sepharose gel, X represents -CH-CN(OH)-CH2- or m is equal to 2,4,6 or n is 2-9, and -NH-SPA is aligand of the protein A. The ligand of the protein A and the sepharose gel microspheres in the adsorption material are connected through ether bonds, the protein A adsorption material is stable in structure, and an adsorbent is stable in color, low in falling amount and high in safety, so that the protein A adsorption material is high in adsorption efficiency and good in regenerability.

The invention relates to a gene engineering antibody technique and a molecular cloning technique and in particular relates to an aflatoxin B1 nano antibody immune adsorption material which is an agarose gel coupled with an aflatoxin B1 nano antibody fusion protein. The amino acid sequence of the aflatoxin B1 nano antibody fusion protein is shown in SEQ ID NO.1, and the nucleotide sequence of the protein is shown in SEQ ID NO.2. A gene engineering technique is adopted, a single-domain heavy chain antibody specifically aiming at aflatoxin B1 is fused with a self-coupled tag protein, then purification-free specific one-step orientated coupling of a recombinant expression ligand can be achieved, preparation procedures of an affine adsorption material can be greatly simplified, the cost can belowered, and meanwhile, occupation of a ligand combination site of a conventional coupling method is avoided, so that the adsorption capacity of an immune affine adsorption material can be increased.

The invention discloses a sepharose gel plate punching structure. The punching structure comprises a supporting stand, a lifting unit, and a plurality of needle tubes; the supporting stand comprises atop plate, a supporting plate and a bottom plate which are arranged in sequence from top to bottom; the bottom plate is provided with a placing surface for placing a sepharose gel plate; the liftingunit comprises at least one supporting plate lifting assembly and at least one top plate lifting assembly, the supporting plate lifting assembly drives the supporting plate to go up and down and the top plate lifting assembly drives the top plate to go up and down; negative pressure assemblies are arranged inside the needle tubes; and each of the negative pressure assemblies comprises a piston which moves up and down inside a punching tube and a piston rod with the lower end connected with the piston, the upper end of the piston rod penetrates through the punching tube and is fixed with the top plate, a through hole is formed in the upper end of the punching tube and is for the piston rod to penetrate, and the needle tube is fixed with the supporting plate. The punching structure can meetactual requirements of different experiments and rapidly punch needed holes.