52 results about "Immobilized protein" patented technology

A method for the preparation of an immobilized protein ultrathin film reactor by immersing a solid support alternately into an aqueous solution of protein, and into an aqueous solution of polyion charged oppositely to said protein and preparing a structurally controlled ultrathin film of mono- or multi- protein layers on a said solid support with precision at molecular level. And a method for chemical reaction of a substrate by preparing an immobilized protein ultrathin film reactor composed of multiple layers of protein on a solid support by above mentioned method, and initiating a chemical change of the substrate molecules using obtained immobilized protein ultrathin film reactor.

Provided are a support for affinity chromatography which has excellent alkali resistance, and a method for isolating immunoglobulin. A support for affinity chromatography, containing an immobilized protein ligand represented by the following formula (1):R—R2  (1)wherein R represents a polypeptide consisting of 4 to 30 amino acid residues that contains an amino acid sequence represented by ATK or ASK; and R2 represents a polypeptide consisting of 50 to 500 amino acid residues containing an immunoglobulin-binding domain consisting of an amino acid sequence represented by SEQ ID NO: 1 or SEQ ID NO: 2, the partial sequence thereof, or an amino acid sequence having 70% or more identity to these sequences; with the proviso that a terminus at which R2 binds to R is C-terminus or N-terminus of the immunoglobulin-binding domain.

The invention belongs to the field of immobilization of proteins, and relates to preparation and application of a dopamine functional magnetic nano-carrier. The preparation comprises the following steps: firstly, dissolving FeSO4.4H2O and FeCl3.6H2O into distilled water, high-speed stirring and under protection of nitrogen, adding ammonia water of 25% NH4OH, and adding oleic acid; secondly, repeatedly cleaning black precipitates obtained by a stirring reaction and then using a strong magnet to perform precipitation separation, and drying to obtain oleic acid magnetic nano-particles; and finally, adding the oleic acid magnetic nano-particles into a dopamine hydrochloric acid solution, after the reaction is over, using a magnet to separate, cleaning and drying obtained deep brown precipitates, and then the dopamine functional magnetic nano-carrier can be obtained. According to the preparation and application of the dopamine functional magnetic nano-carrier provided by the invention, dopamine hydrochloride is adopted for modifying magnetic oleic acid nano-particles, so that the surface of a magnetic nano-particle has a plentiful of quinonyls and amidogen functional groups, thus proteins can be effectively immobilized; besides, the surface of the carrier has a layer of soft poly dopamine oxide films, the surface area is larger, and the preparation method is simple.

The present embodiments provide systems, kits and methods suitable for performing dry or substantially dry electro-blotting analyses on immobilized protein or nucleic acid samples. Electro-blotting performed according to the presently described embodiments may include a step whereby detection of one or more immobilized proteins or nucleic acids is electrophoretically accelerated. Methods for performing electro-blotting of immobilized proteins or nucleic acids may include applying an electric voltage to one or more reagents typically used in protein or nucleic acid blotting procedure. The one or more reagents may be absorbed on a suitable carrier matrix. Electro-blotting performed in accordance with the systems and methods described herein may be performed under substantially dry conditions (i.e., with little or no aqueous buffers).

The invention discloses a graphene (GR)-hydrotalcite-like compound (LDH)-graphite phase carbon nitride (g-C3N4) composite material immobilized protein modified electrode, a production method and an application thereof. The method comprises the following steps: compounding an exfoliation hydrotalcite-like compound with negatively charged exfoliation graphene oxide nanosheets, mixing the obtained material with melamine, roasting, and adding water to recover in order to prepare a GR-LDH-g-C3N4 hybrid; and adding a GR-LDH-g-C3N4 and Hb mixed solution to an ionic liquid modified carbon paste electrode in a dropwise manner in order to produce the GR-LDH-g-C3N4 composite material immobilized protein modified electrode. The graphene-hydrotalcite-like compound-graphite phase carbon nitride composite material immobilized protein modified electrode fully displays advantages of a gel method and an integrating, merging and layering method, increases the specific surface area, the conductivity and the biocompatibility of the hybrid material, and inhibits the afresh stacking of GR sheets; and a roasting method allows g-C3N4 to grow on the surface of a GR sheet layer, increases the specific surface area of the hybrid material, and improves the electron transfer rate of an electrochemical reaction process. A constructed CTS / GR-LDH-g-C3N4-Hb composite film-based third-generation trichloroacetic acid sensor has the advantages of low detection limit, wide detection range and small Michaelis constant.

A method for purifying a protein using Protein A chromatography comprising a) absorbing the protein to Protein A immobilized on a solid support; b) removing contaminants by washing the immobilized Protein A containing the absorbed protein with a buffer comprising one or more chaotropic agents in combination with one or more hydrophobic modifiers and having a pH of at least 7.0; and c) eluting the protein from the Protein A immobilized on the solid support.

The invention discloses a method for purifying and directionally immobilizing histidine tag protein and application. According to the method disclosed by the invention, tannic acid, metal ions and a hydrophilic anti-protein adhesion polymer are self-assembled and modified on the surface of a carrier to obtain a material surface which is efficiently chelated with cobalt ions, so that one-step purification and immobilization of the histidine tag protein is realized; a ligand of trivalent cobalt has inertia and divalent cobalt ions are oxidized into trivalent cobalt ions so that the stability of the immobilized protein can be improved. The method disclosed by the invention has the advantages of simple process and low price; a separation and purification or immobilization operation process for the histidine tag protein of the method is simple and effective, so that the technology has a wide application prospect.

Methods and reagents for site-selective functionalization of peptides and proteins. The methods most generally involve the reaction of a thioester with hydrazine. Reagents include bifunctional reagents of formula:H2N—NH—CH2-M-L-FGand salts thereof where M is a single bond or a chemical group carrying a non-bonding electron pair, such as —C(O)NR′—, where R′ is H, or an alkyl or aryl group; L is an optional linker group as described above; and FG is a functional group having reactivity that is orthongonal to that of the hydrazine group. FG can, among others, be an azide, alkenyl, alkynyl, nitrile (—CN) or triazole group and is preferably an azide group (—N3). Methods and reagents can, for example, be combined with intein-mediated protein splicing to link proteins or fragments thereof to various chemical species or to a surface. Surface immobilization of proteins via the methods herein results in immobilized proteins which substantially retain biological activity and is thus useful for the generation of peptide or protein microarrays. Kits for functionalization and / or immobilization of peptides and proteins are provided as well as microarrays of peptides, proteins or both.

An immobilized protein catalyst is prepared by applying an adhesive to a polymeric support, applying a layer of a globular protein over the layer of adhesive, binding a crosslinking agent to the protein layer, and binding the protein catalyst by reaction with the crosslinking agent.

The invention discloses preparation and electrochemical sensing application research of a tin dioxide-three-dimensional graphene (SnO2-3DGR) nanocomposite immobilized protein modified electrode, and belongs to the technical field of biosensors. A method provided by the invention comprises the following steps: preparing a SnO2-3DGR nanometer material, dispensing the SnO2-3DGR nanometer material onto the surface of a carbon ionic liquid electrode (CILE), then dispensing myoglobin (Mb) onto the SnO2-3DGR modified CILE surface, naturally airing, dropwise adding a chitosan (CTS) solution, and drying a third-generation electrochemical enzyme sensor. The prepared sensor has the characteristics of a simple process, convenient operation, low cost, high detection sensitivity, strong electrical signal and simple electrode pretreatment, and can be used for detecting organic small molecular trichloroacetic acid (TCA), with a detection limit of 1.67 mmol / L.

The present invention reates to a method for detecting activated ras protein. The method includes immobilizing a protein on a solid support, incubating the immobilized protein with lysates from cultured cells, where the lysates include activated ras protein, and determining the amount of activated ras protein bound to the immobilized protein. The present invention also relates to a method of detecting ras oncogenic related malignancy in a human subject.

The invention discloses a method for screening and characterizing a metabolite-protein interaction system based on metabolomics and a structural mass spectrometric technique; a heterologously expressed tagged target protein is immobilized through an affinity immobilization method, then, the target protein is incubated with a metabolite extract, and finally, after being separated, denatured and eluted, the target protein-metabolite complex is subjected to non-targeted metabolomics analysis, and the complex is compared with a control group (an affinity vector without immobilized protein) so as to screen a metabolite with potential interaction with the target protein. Afterwards, the potential interaction system is directly verified using structural mass spectrometry. The binding method enables high-throughput screening of metabolites with potential interaction with the target protein, and also, direct characterization of the structural mass spectrometry can be used to screen out false positive interaction results that are probably introduced during indirect characterization. The method is a reliable high-throughput research tool for protein-metabolite interactions.

The invention relates to the technical field of gene engineering, in particular to CD86-based membrane fixed protein, and a preparation method and application thereof. CD86 comprises a signal peptide encoding gene, an extracellular region encoding gene, a transmembrane region encoding gene and an intracellular region encoding gene; and the membrane fixed protein is to replace the extracellular region encoding gene of the CD86 with an expressible exogenous gene. Lentivirus-containing host cells can be used for cultivating cells, and can supply nutritional substances for the cells continuously, additional nutritional substances are not needed, and convenience and rapidness are realized.

Provided are a support for affinity chromatography which has excellent alkali resistance, and a method for isolating immunoglobulin. A support for affinity chromatography, containing an immobilized protein ligand represented by the following formula (1):R—R2  (1)wherein R represents a polypeptide consisting of 4 to 30 amino acid residues that contains an amino acid sequence represented by ATK or ASK; and R2 represents a polypeptide consisting of 50 to 500 amino acid residues containing an immunoglobulin-binding domain consisting of an amino acid sequence represented by SEQ ID NO: 1 or SEQ ID NO: 2, the partial sequence thereof, or an amino acid sequence having 70% or more identity to these sequences; with the proviso that a terminus at which R2 binds to R is C-terminus or N-terminus of the immunoglobulin-binding domain.

The invention relates to a method for screening active ingredients of traditional Chinese medicine by nano mesoporous material immobilized target protein. The method includes filling mesoporous material immobilized protein into a chromatographic column, preparing an immobilized target protein inhibitor screening model, dissolving inhibitor having representative inhibiting effect on the immobilized protein into buffer solution to serve as a mobile phase, introducing samples of substrates with saturation concentration every fixed time, collecting peak appearance fraction, replacing the mobile phase with buffer solution free of inhibitor after fixed times, introducing samples of the substrates with saturation concentration every fixed time, respectively freezing and drying the fraction, dissolving the fraction into water, separating the substrates from a product through the analytical type chromatographic column, recording peak area of the product, drawing an inhibition curve of the immobilized target protein inhibitor, checking the model and adopting the prepared model to dissolve aqueous extract of traditional Chinese medicine to be screened into the buffer solution to serve as the mobile phase. Whether the traditional Chinese medicine has inhibition effect or not and the process and the type of the traditional Chinese medicine can be judged according to the peak area of the product. The method has the advantages of high reliability, online performance and the like.

The present invention provides a method for inexpensively, easily and efficiently modifying the C-terminus of a protein or peptide; a method for easily and reliably isolating a C-terminal peptide fragment of a protein or peptide; and a method for rapidly, accurately and reliably determining an amino acid sequence of a protein or peptide by using a mass spectroscope. A method comprising the step of adding a formylation reagent and a catalyst to a protein or peptide to convert a carboxyl group into an aldehyde group. A method comprising the step of reacting a nucleophilic reagent with the aldehyde group to modify the C-terminus of the protein or peptide. A method comprising the step of reacting a support having a nucleophilic group to immobilize the protein or peptide. A method comprising the step of fragmenting the immobilized protein or peptide, washing the support, and isolating the C-terminal peptide fragment from the support. A method comprising the step of subjecting the isolated C-terminal peptide fragment to mass spectrometry and determining an amino acid sequence.

It is an object of the present invention to provide a method of conveniently efficiently and easily immobilizing a protein on a substrate, a method of detecting a substance capable of interacting with the protein at a high sensitivity, and a protein and a protein-immobilized substrate which are used in these methods. In the method of detecting a substance capable of interacting with a protein using the immobilized protein of the present invention, a protein bound to a compound having a group capable of binding onto a substrate for immobilizing a biomolecule or a carrier provided on the substrate and immobilized onto the substrate, is used as the immobilized protein.

The invention provides a method for forming an immobilized agent gradient within a 3-dimensional porous scaffold. A 3-dimensional scaffold formed from a biocompatible material is provided. The surface of the scaffold and / or the agent is activated so as to allow binding of the agent to the scaffold. The activated scaffold is contacted with a solution containing the agent. Contact with the solution is maintained for a sufficient period of time to allow diffusion of the solution through a portion of the scaffold, thereby forming a desired gradient of the agent through the 3-dimensional scaffold.

The present invention provides a method for detecting modified LDL, abnormal cells or bacteria using an intermolecular interaction analysis method, in which a region involved in ligand recognition by a receptor is expressed, without modification or as a biotinylated protein, in cells or in a test tube, and thereafter, the expressed region or the expressed biotinylated protein is immobilized via avidin or streptavidin to a solid phase while the orientation thereof is maintained, and the immobilized protein is utilized; and a kit for detecting the modified LDL or the like.

The invention provides methods to assess protein stability and to obtain sizing information. In one aspect, the screen comprises a 94 detergent panel and a series of MWCO filtered microplates. A protein of interest is bound to an affinity matrix and aliquoted into a 96-well microplate. Wells containing the immobilized protein are washed in the new detergent and then eluted in the new detergent into a collection plate. Protein not stable in the new detergent is precipitated on the resin and not present in the elutions. Half of the elution is passed through a high (i.e., 300 kDa) MWCO microplate and the other half through a low (i.e., 100 kDa) MWCO microplate. Elutions from the microplates are spotted on a nitrocellulose membrane, visualized by Western analysis (or by some other method), and quantified. The high MWCO provides stability readout and the ratio of low / high kDa provides sizing information.

The invention belongs to the field of protein immobilization and discloses preparation of a functionalized graphene oxide carrier and an application method of the functionalized graphene oxide carrier in rapid immobilization of proteins. According to the technical scheme, the invention has the key points of preparing and modifying functionalized graphene oxide; P-beta-sulfatoethylsulfonyl aniline (SESA) is adopted to activate the graphene oxide (GO), diazotization is carried out on the activated carrier, and finally, the carrier is used for rapidly immobilizing the proteins. The functionalized graphene oxide carrier has a lamellar structure and is large in surface area and simple in preparation method. According to the preparation method and the application provided by the invention, three different proteins in different shapes are used for immobilization, the immobilization can be rapidly finished with the functionalized graphene oxide as an immobilization carrier, and in addition, the immobilization yield and the immobilization efficiency can both reach 80-90 percent. Therefore, the method of preparing the functionalized graphene oxide carrier for immobilizing the proteins provides a new thinking to study rapid immobilization and has certain study significance.

This invention provides a process for effectively producing an immobilized protein that is immobilized only at the carboxy terminus. This is a process for immobilizing on a carrier for immobilization a protein represented by general formula (1):NH2—R1—COOH   (1)wherein a protein having a sulfhydryl group represented by general formula (2) is produced:NH2—R1—CONH—R2—CO—NH—CH(CH2—SH)—CO—NH—R3—COOH   (2),the resulting protein is immobilized adsorptively on the carrier for immobilization represented by general formula (3) via ion interactions under neutral conditions:NH2—Y   (3), anda sulfhydryl group of the cysteine residue in the protein represented by general formula (2) that is adsorbed on the carrier for immobilization represented by general formula (3) is cyanated using a cyanating reagent to convert it into a cyanocysteine residue,thereby producing an immobilized protein represented by general formula (4):NH2—R1—CO—NH—R2—CO—NH—Y   (4)wherein R1 and R2 represent arbitrary amino acid sequences, R3 represents an amino acid sequence that has a strong negative charge around the neutral region and can acidify the isoelectric point of NH2—R1—CONH—R2—CO—NH—CH(CH2—SH)—CO—NH—R3—COOH, and Y represents an arbitrary carrier for immobilization.

The invention relates to the field of immobilized protein, in particular to a method for preparing heterologous protein wrapped polyhedrosis. The bombyx mori cytoplasmic polyhedrosis protein, a fusion foreign protein with a bombyx mori cytoplasmic polyhedrosis virus structural protein VP5 tag and a protease specific cleavage site are co-expressed in bombyx mori culture cells or bombyx mori by using a baculovirus expression system through a genetic engineering method, so that the polyhedrosis wrapping the foreign protein is formed. The formed polyhedron can be purified through simple differential centrifugation; the polyhedrin has protection and slow release effects on foreign proteins wrapped by the polyhedrin. After the purified polyhedrosis is cracked under an alkaline condition, the pH value is adjusted to a polyhedrosis protein isoelectric point by using a hydrochloric acid solution, the polyhedrosis protein can be precipitated through centrifugation, and the fusion foreign protein is retained in a supernatant, so that the fusion foreign protein can be quickly and conveniently obtained; the fusion foreign protein is subjected to enzyme digestion through corresponding protease to remove a VP5 tag, so that a foreign protein closer to the nature can be conveniently obtained.

The invention discloses a method for immobilizing a protein by using a biofilm. The microbial biofilm is used as an initial material, and chemical modification with a polysaccharide is performed to obtain a biofilm material for immobilization of a target object, wherein the target object comprises a free amino-containing protein and / or an amino-containing nanoparticle. The biofilm material disclosed by the invention can be used to immobilize proteins with catalytic activity or non-catalytic activity, and free amino-containing nanoparticles, and the stability of the immobilized protein can besignificantly improved. In addition, the method is universal and can be extended into the use of universal type biofilms, and the utilization range of biofilm materials is greatly improved.