485 results about "Antibody Affinity Chromatography" patented technology

The invention relates to the technical field of biology, and particularly discloses a recombinant collagen III and application thereof in preparation of gel. The recombinant collagen III is designed by optimizing functional area sequences of human I-type and III-type collagen, the amino acid sequence is shown as SEQ ID No.2, and the recombinant collagen III has the characteristics of high stability, good hydrophilicity and low immunogenicity. The preparation method comprises the steps of expression vector construction, escherichia coli induced expression, affinity chromatography purification and renaturation. The recombinant collagen III can be prepared into a gel dressing and comprises sodium alginate, methylparaben and other components. Experiments show that the gel can effectively promote cell proliferation and has no cytotoxicity; in a mouse skin injury model, the collagen can relieve inflammation, accelerate wound healing and inhibit scar formation, and the effect of the collagen is superior to that of natural human III-type collagen. The invention provides a safe and efficient novel material for wound repair, and is suitable for medical dressings and tissue engineering.

The embodiment of the invention discloses human-derived recombinant III-type collagen as well as a preparation method and application thereof. The amino acid sequence of the human-derived recombinant III-type collagen comprises a basic repetitive unit, and the amino acid sequence of the basic repetitive unit is as shown in SEQ ID NO.4. The amino acid sequence of the basic repetitive unit is as shown in SEQ ID NO.3. The amino acid sequence of the basic repetitive unit is as shown in SEQ ID NO.4. The amino acid sequence of the human recombinant III type collagen is shown as SEQ ID NO.3. The invention also discloses a preparation method of the human recombinant III type collagen. According to the invention, a human recombinant III-type collagen gene with the coding molecular weight of 60kD is designed, an escherichia coli expression strain is constructed and expressed, the yield of the human recombinant III-type collagen is increased through amplification culture on a fermentation tank, and the protein purity is improved by virtue of strategies such as ultrafiltration primary purification and affinity chromatography. The human-derived recombinant III-type collagen prepared by the invention avoids potential hidden dangers such as animal diseases or viruses and immunological rejection caused by a traditional extraction method, has excellent medical performance, can effectively control the production cost and the product quality by virtue of a fermentation preparation process, and is also easy to realize large-scale production.

The invention relates to a genetically engineered bacterium for producing a resuscitation promoting factor Rpf and application of the genetically engineered bacterium, and belongs to the technical field of bioengineering. The genetically engineered bacterium is a prokaryotic cell, and the nucleotide sequence of the resuscitation promoting factor Rpf is as shown in SEQ. ID. NO.1. The genetically engineered bacterium is preserved in China Center for Type Culture Collection. The Rpf recombinant protein prepared from the genetically engineered bacterium constructed by the invention is in soluble expression, and a good research foundation is laid for subsequent detection of VBNC state pathogenic bacteria. After the recombinant protein is purified through affinity chromatography, the purity of the Rpf recombinant protein can reach 93% and is far higher than the extraction purity in the prior art.

The invention discloses recombinant silk fibroin peptide as well as a preparation method and application thereof, and belongs to the technical field of cosmetic raw material development. The amino acid sequence of the recombinant silk fibroin peptide SFHR6 disclosed by the invention is as shown in SEQ ID NO. 1. Soluble expression of the recombinant silk fibroin peptide is achieved through prokaryotic expression and eukaryotic expression, the high-purity soluble recombinant silk fibroin peptide SFHR6 is obtained through nickel column affinity chromatography, hydrophobic chromatography, cation chromatography and anion chromatography technologies, and the recombinant silk fibroin peptide SFHR6 is high in purity, high in tyrosinase inhibition rate and strong in whitening effect. The purification process provided by the invention is simple and convenient, has good repeatability, effectively solves the quality control problem of naturally extracted protein products, has the advantages of high efficiency, high-purity heterologous expression and low large-scale production cost, and has relatively high industrial application potential.

The application discloses a method for recovering active glycoprotein Patatin from potato starch processing waste juice, and belongs to the technical field of protein extraction. The method is characterized in that the active Patatin protein is efficiently separated by using the combination of ammonium sulfate fractionation precipitation, anion exchange chromatography, Con A affinity chromatography and gel chromatography, and the obtained Patatin protein retains the original protein structure and biological function activity. Compared with the prior art, the method has better separation purity and higher efficiency, and the obtained Patatin protein has complete protein higher structure and function activity, and has a better application scene.

The invention relates to the technical field of biological medicine, in particular to a preparation method and application of a function-enhanced mesenchymal stem cell exosome, the preparation method comprises the following steps: S1, preparing an NK cell exosome, collecting supernate in an NK cell culture medium, and treating the supernate by using an optimized exosome separation and purification process to obtain an NK cell exosome; finishing preparation of a stock solution; s2, culturing the mesenchymal stem cells, culturing the cells in a 3D suspension culture mode, treating and controlling low oxygen concentration by combining with NK cell exosomes, and simulating an in-vivo inflammation microenvironment in a multi-azimuth manner, so that the exosomes secreted by the mesenchymal stem cells have a stronger immune regulation function and an anti-inflammatory effect; s3, preparation of mesenchymal stem cell exosomes: preparing a stock solution through an exosome separation and purification process combining a tangential flow ultrafiltration technology and a PS affinity chromatography technology; the exosome stock solution obtained by the method is high in purity and high in yield, and has more excellent biological activity and excellent treatment effect.

The invention discloses a method for separating and purifying bovine lactoferrin by using a ligand affinity chromatography technology, and belongs to the technical field of protein purification. The method comprises the following steps: introducing a bovine lactoferrin gene into pichia pastoris, fermenting to obtain a pichia pastoris fermentation solution, centrifuging, resuspending, homogenizing at high pressure, centrifuging and filtering the pichia pastoris fermentation solution to obtain a filtrate, separating and purifying bovine lactoferrin in the filtrate by using an affinity chromatography technology, immobilizing a triazine dye ligand in a filler of a chromatographic column, and purifying the triazine dye ligand to obtain the bovine lactoferrin. An eluent is prepared, the concentration of NaSCN in the eluent is 0.2-0.5 mol / L, the volume concentration of ethylene glycol in the eluent is 15%-20%, the concentration of NaCl in the eluent is 1-2 mol / L, and the bovine lactoferrin in the chromatographic column is eluted. According to the method, the composition of the eluent is optimized, the three components in the eluent are matched with one another and have a synergistic effect, the stability of the bovine lactoferrin is protected, the purity of the bovine lactoferrin is guaranteed, meanwhile, the elution efficiency of the bovine lactoferrin is improved, and the dosage of the eluent is reduced.

The invention provides a bispecific antibody and a purification method thereof. The purification method comprises the following steps: a) loading a sample containing a bispecific antibody to a affinity chromatographic column, and leaching and eluting; the biomacromolecules in the sample are adsorbed on the affinity chromatography column in the leaching stage, the biomacromolecules comprise antibodies, host cell proteins and protein aggregates, and the antibodies comprise bispecific antibodies and homodimers; separating the antibody from the affinity chromatography column in the elution stage, and collecting the eluate in the elution stage; b) loading the eluent to a hydrophobic membrane material, carrying out flow-through, and collecting a flow-through liquid containing the purified antibody; and the flow-through liquid used for flow-through contains 0.73 to 0.77 M of (NH4) 2SO4. The method can solve the problem that homodimers in bispecific antibodies are difficult to efficiently remove in the prior art, and is suitable for the field of antibody purification.

The present invention provides a heparin affinity chromatography medium and a preparation method thereof. The heparin affinity chromatography medium comprises a porous polymer microsphere matrix modified with a polyamide-amine dendrimer to obtain a modified matrix; heparin is coupled to the modified matrix to obtain the heparin affinity chromatography medium. The heparin affinity chromatography medium of the present invention has a high adsorption capacity for target biomolecules, significantly improving separation efficiency; and also exhibits high mechanical strength.

The invention discloses a method for purifying sialic acid from fermentation liquor, and belongs to the field of biochemical engineering separation and purification. The method comprises the following steps: firstly, concentrating sialic acid biological fermentation liquor by adopting membrane separation, adjusting the pH value of the concentrated liquor to 7-11, loading a sample on a boric acid affinity chromatography column, respectively washing with water and acid water, and collecting a sialic acid eluate; adjusting the pH value of the saliva acid eluate to 5-10, loading the saliva acid eluate to anion exchange resin, washing with water and acid water to obtain a resin desorption solution, and adding the resin desorption solution into a spray dryer for drying to obtain a sialic acid finished product. The stable five-membered ring complex is generated by utilizing the action of a boric acid group and a dihydroxyl structure in sialic acid. According to the method, the sialic acid is subjected to complexing under an acidic condition, molecules are eluted from a medium, selective separation of the sialic acid is realized, the sialic acid is further refined and purified by adopting anion exchange resin, high-purity sialic acid is prepared, introduction of a crystallization process is avoided, and use of an organic solvent and an acid reagent is greatly reduced.

The invention discloses an influenza C virus HEF protein polyclonal antibody preparation method, which comprises: carrying out signal peptide prediction, antigen epitope screening and hydrophobicity analysis through bioinformatics, constructing a recombinant prokaryotic expression vector pET32a-HEF by using a seamless cloning technology, and efficiently expressing the HEF protein in Escherichia coli to obtain the influenza C virus HEF protein polyclonal antibody. The inclusion body protein yield is improved by optimizing conditions; and purifying by nickel column affinity chromatography to obtain high-purity recombinant protein, emulsifying the high-purity recombinant protein and a Freund's adjuvant, immunizing a Japanese white rabbit, immunizing for three times, and collecting high-titer serum. The method breaks through detection limitation caused by antigen tag concealment in a eukaryotic expression system, the obtained polyclonal antibody is high in specificity, the eukaryotic expression HEF protein can be accurately recognized, and a key tool is provided for ICV virus-like particle identification, vaccine research and development and diagnosis technologies. Compared with a traditional method, the scheme is simple and convenient to operate, low in cost and high in antibody titer, and has remarkable application value.

The invention relates to the technical field of biology, in particular to a method for producing vitamin B12 through fermentation of propionibacterium shermangii, which comprises the following steps: inoculating propionibacterium shermangii into a seed culture medium to culture a seed solution; inoculating the seed solution into a fermentation culture medium for fermentation; after fermentation is finished, thalli are removed in a centrifugal separation or filtration mode, a fermentation solution is subjected to ultrafiltration concentration, a concentrated solution is purified through ion exchange chromatography or affinity chromatography, and a vitamin B12 crude product is obtained; and further purifying by adopting a crystallization method to obtain a high-purity vitamin B12 finished product. According to the method for producing the vitamin B12 through fermentation of the propionibacterium shermannii, the yield and the purity of the vitamin B12 are remarkably improved and the production cost is reduced by accurately controlling fermentation conditions, optimizing culture medium components, introducing an intelligent regulation and control system and adopting a specific purification process, so that the method is suitable for industrial production. The problems of low yield, unstable purity, difficult production process control and the like of the vitamin B12 in the prior art are solved.

The invention relates to the technical field of protease separation, in particular to a novel earthworm protease separation and purification process. According to the present invention, the ultra-high speed separation, the molecular weight spectrum comparison extraction and the affinity chromatography purification scheme are adopted, such that the different biological macromolecules in the earthworm can be effectively separated, the earthworm protease-containing cell lysis solution can be concentrated, and most of the impurities can be removed so as to obtain the high-purity protease solution, the method is simple in operation and does not depend on complex chemical reactions, contact between protein and chemical reagents can be reduced, meanwhile, damage of high temperature to earthworm protease is avoided through a low-temperature centrifugation scheme, the enzymatic activity of earthworm protease is kept, high-purity and high-activity earthworm protease is obtained through affinity chromatography purification, operation is easy, and the method is suitable for industrial production.

The invention discloses preparation and application of grouper enterospora sporowall protein SWP26 and a polyclonal antibody of the grouper enterospora sporowall protein SWP26, and belongs to the field of animal quarantine, the grouper enterospora sporowall protein SWP26 is obtained by amplifying grouper enterospora genes by using a PCR (Polymerase Chain Reaction) method, shearing a target band and transferring the target band into a pET-32a vector to construct a recombinant plasmid, and then transforming escherichia coli BL21 (DE3) for induced expression. A Ni-NTA affinity chromatography method is used for protein purification, animal immunization and polyclonal antibody purification are carried out on the purified SWP26 recombinant protein, western blot detection is carried out on the purified antibody, an obvious signal appears at about 43kDa, and it is proved that the anti-SWP26 polyclonal antibody can be subjected to a specific reaction with the SWP26 purified protein. The invention clones and identifies the high-abundance spore wall protein SWP26 positioned on the surface of the enterosporidium of the grouper for the first time, belongs to a specific protein of the enterosporidium of the grouper, and can be used as a drug target for treating enterocytozoonosis of the grouper.

The invention relates to the technical field of biological pharmacy, in particular to a high-yield mutant streptococcus protein G and application thereof. Compared with a wild type protein G, the mutant recombinant protein G provided by the invention has the advantages that the yield in an expression system in escherichia coli is multiplied, the cost for preparing a G Column filler is greatly reduced, and the problem of high cost of a monoclonal antibody is solved. The mutant recombinant protein G retains the binding function of wild protein G to IgG protein, can greatly reduce the manufacturing cost of a G Column affinity chromatography column, is more stable and not easy to degrade, has more recycling times while reducing the production cost of the protein G affinity chromatography column, and has wide application prospects in an antibody purification process.

The invention discloses a cod bone-derived high-activity calcium chelating peptide and a preparation method thereof, and belongs to the technical field of functional biological product preparation. The calcium chelating peptide provided by the invention is derived from a cod bone collagen enzymatic hydrolysate and is obtained through affinity chromatography separation of hydroxyapatite, the calcium chelating activity is 4.9-5.2 mu g / mg, and the calcium chelating peptide contains aspartic acid and glutamic acid enriched peptide fragments, through deep processing of cod bones, high-valued utilization of cod bone resources is realized, the high-activity calcium chelating peptide is prepared, and the calcium chelating peptide has a wide application prospect. And a high-quality raw material is provided for the development of a novel calcium supplement.

The invention discloses a preparation method and application of a tetravalent hand-foot-and-mouth disease subunit-new adjuvant vaccine, and relates to the technical field of hand-foot-and-mouth disease vaccines. VP1 proteins of EV-A71, CV-A16, CV-A10 and CV-A6 are expressed through an insect cell-baculovirus system, the VP1 proteins are purified through a nickel affinity chromatography column and mixed with an MF59 adjuvant to prepare a tetravalent vaccine, in-vivo evaluation of mice is conducted, and meanwhile the immunogenicity and protection efficacy of the tetravalent VP1 protein vaccine and the immunogenicity and protection efficacy of a monovalent VP1 protein vaccine are compared. Immune results show that compared with a VP1 monovalent vaccine, all serum of a mouse inoculated with the tetravalent VP1 protein vaccine shows strong reactivity to all captured antigens. Therefore, the results show that the prepared tetravalent subunit-new adjuvant vaccine is a promising broad-spectrum vaccine.

The invention provides a preparation method of thermal-stability T7RNA polymerase and the thermal-stability T7RNA polymerase, belongs to the field of biology, and can solve the problems that an existing T7RNA polymerase preparation process is complex, the expression culture period is long, multi-step purification treatment is needed, liquid change treatment needs to be carried out independently before storage, and the production cost is high. The method comprises the following steps: transferring plasmids containing a T7RNA polymerase gene sequence into expression host bacteria M15, culturing, screening, cloning, adding glycerol for strain preservation, and establishing a strain three-level library; the method comprises the following steps: taking glycerol bacteria, carrying out streak resuscitation on a flat plate, selecting monoclone, inoculating the monoclone into an LB culture medium, culturing, carrying out enlarged culture, carrying out IPTG induced expression, centrifugally collecting thalli, cleaning, adding a lysis buffer solution, resuspending, carrying out ultrasonic cell disruption, centrifuging, and collecting supernate; purifying by nickel column affinity chromatography, removing impure protein by an impure washing buffer solution, eluting target protein, purifying by matching with anions and cations, and collecting the purified protein; and mixing the purified protein with a storage buffer solution, and subpackaging and storing. The method can be applied to rapid preparation of the thermal-stability T7RNA polymerase.

The invention provides a method for screening a hyaluronidase (HAase) inhibitor, and belongs to the technical field of medicines. The method comprises the following steps: firstly, rapidly identifying a natural product with HAase inhibition activity by adopting a fluorescence sensing technology according to the principle that carbon quantum dots (CDs) are used as a fluorescence probe, hyaluronic acid (HA) modified gold nanoparticles are used as a quencher, and the fluorescence of the CDs can be quenched by the quencher; when HAase exists, HA can be enzymatically decomposed to release CDs, and fluorescence recovery is caused; when a screened sample has HAase inhibition activity, CDs fluorescence recovery can be prevented. And 2, capturing components in a sample capable of inhibiting HAase activity by virtue of an affinity chromatography technology, immobilizing HAase on a carrier material by virtue of a method, eluting by virtue of a solvent to obtain components capable of being combined with HAase, and identifying the components by virtue of a liquid chromatography-mass spectrometry technology. According to the method, the HAase inhibitor can be rapidly and accurately screened from natural products.

The invention discloses an escherichia coli-bacillus subtilis shuttle plasmid and application, and belongs to the field of gene engineering. The shuttle plasmid comprises the following elements: a promoter sequence, a coding sequence of bacillus subtilis signal peptide, a replication initiation sequence of bacillus subtilis, a replication initiation sequence of escherichia coli and an antibiotic resistance gene. According to the plasmid design, a modular structure is adopted, and quick replacement of key elements such as a promoter, a signal peptide and RBS is supported through optimized enzyme cutting site layout; a 6 * His tag introduced at the tail end of a multiple cloning site (MCS) can be adapted to nickel column affinity chromatography, and the purification process of the recombinant protein is simplified. The plasmid has the characteristics of small molecular weight and convenience in operation, can be efficiently cloned and constructed in escherichia coli and then directly converted into bacillus subtilis for protein expression, is particularly suitable for development of a gram-positive bacterium secretory expression system, and provides an efficient tool for large-scale production of industrial enzyme preparations.

The current invention reports a method for purifying an antibody by reducing the content of a host cell protein. The method employs a wash step with a low conductivity aqueous solution in an affinity chromatography.

The invention relates to the technical field of agricultural fertilizers. More specifically, the invention relates to an extraction method of tenebrio molitor antibacterial peptide and application of the tenebrio molitor antibacterial peptide in preparation of tea tree anthracnose resistant fertilizer. According to the preparation method, the colletotrichum camelliae liquid is taken as an induction factor, the ultrasonic induction technology is utilized to strengthen the absorption of tenebrio molitor to the induction factor, meanwhile, a supercritical / ethanol cosolvent extraction process and a bionic affinity chromatography purification process are combined, the efficient extraction of the antibacterial peptide is realized, and the obtained antibacterial peptide has a relatively good inhibition effect on the colletotrichum camelliae. A tea tree anthracnose resisting leaf fertilizer is further prepared through the tenebrio molitor antibacterial peptide, the leaf fertilizer has a good prevention and treatment effect on tea tree anthracnose caused by camellia colletotrichum spinosus, meanwhile, tea growth can be promoted, and the yield reduction caused by anthracnose and nutrition deficiency of tea tree crops can be effectively reduced.

The invention discloses a PD-L1 / TGF-beta / STING triple targeting antibody coupling drug as well as a preparation method and application thereof, and belongs to the field of tumor immunotherapy. The ADC structure is Ab-[L-D] n, Ab is a bispecific antibody YM101 targeting PD-L1 and TGF-beta, L is a cleavable connexon, and D is an STING agonist. The preparation method comprises the following steps: carrier molecule deprotection, linker functionalization, linker-load coupling, antibody sulfhydrylation modification, coupling reaction and affinity chromatography purification. The ADC can increase the number of CXCL16 < + > macrophages, dendritic cells and CXCR6 < + > T cells infiltrated in tumors, effectively reverses the drug resistance of a TGF-beta * PD-L1 bispecific antibody, has a strong anti-tumor effect and no obvious toxic or side effect, and has a good application prospect in preparation of anti-tumor drugs.

The invention discloses a recombinant escherichia coli high-efficiency soluble expression semeglutide 9-37 amino peptide and a purification method, and belongs to the technical field of gene engineering. According to the invention, fusion protein mRNA folding is optimized, hairpin structures are reduced, and efficient soluble expression of fusion protein in escherichia coli is realized. The fusion protein is provided with an SUMO label and an enterokinase cleavage site, the fusion protein is purified through nickel ion affinity chromatography, the fusion protein is subjected to enterokinase specific recognition cleavage and then subjected to nickel ion affinity chromatography and gel filtration chromatography separation, Arg34GLP-1 (9-37) with the purity larger than or equal to 95% is obtained, the wet thallus amount is 150-200 g / L after fermentation culture, and the pure protein yield is 2-3 g / L. The invention provides a fusion expression and rapid purification method of the semeglutide Arg34GLP-1 (9-37) polypeptide, and a solid early-stage foundation is laid for industrial, efficient, green and low-cost preparation of the semeglutide.

The invention belongs to the field of protein purification, and relates to a method for preparing bevacizumab, which comprises the steps of affinity chromatography, virus inactivation, deep filtration, composite anion exchange chromatography, cation exchange chromatography and the like.

The present invention relates to the chromatographic isolation of a target cell or another complex biological material, in particular by column chromatography such as affinity chromatography or gel permeation chromatography. The invention employs a receptor binding reagent that binds to a receptor molecule that is located on the surface of a target cell. The invention in general provides novel methods for the traceless isolation of biologic materials such as cells, cell organelles, viruses and the like. The invention also relates to an apparatus for the isolation of cells and other complex biological materials.

Provided herein are fusion proteins and methods for purifying nucleic acids from a composition. The disclosed compositions and methods allow for faster, more efficient purification of nucleic acids compared to traditional affinity chromatography.

The utility model discloses a novel affinity chromatography column which comprises a chromatography column body, the positioning part comprises side plates, a supporting frame, a supporting disc, a positioning semi-ring, a limiting ring, a sliding block, an inserting ring, an inserting groove, a sliding groove and a drainage tube, the drainage tube is connected to the bottom of the chromatographic column body, the side plates are connected to the lower portions of the two sides of the outer surface of the chromatographic column body, the sliding groove is formed in each side plate, and the sliding blocks are movably connected to the interiors of the sliding grooves; a supporting frame is connected to one side of the outer surface of the sliding block, a positioning semi-ring is connected to the lower portion of one side of the supporting frame, and a supporting disc is connected to the top of the drainage pipe. The utility model relates to an affinity chromatography column, and aims to solve the problems that a drainage tube needs to be mounted at the bottom of the chromatography column to drain extracted fish Igm monoclonal antibodies during actual use of an existing affinity chromatography column, a supporting structure is not arranged between the drainage tube and the chromatography column, the drainage tube and the chromatography column are loosened after long-time drainage, the drainage tube even falls off under serious conditions, and the service life of the chromatography column is influenced. And the stability is poor.