102 results about "Staphylococcal protein" patented technology

The present invention relates to an immunoglobulin-binding protein, wherein at least one asparagine residue has been mutated to an amino acid other than glutamine or aspartic acid, which mutation confers an increased chemical stability at pH-values of up to about 13-14 compared to the parental molecule. The protein can for example be derived from a protein capable of binding to other regions of the immunoglobulin molecule than the complementarity determining regions (CDR), such as protein A, and preferably the B-domain of Staphylococcal protein A. The invention also relates to a matrix for affinity separation, which comprises an immunoglobulin-binding protein as ligand coupled to a solid support, in which protein ligand at least one asparagine residue has been mutated to an amino acid other than glutamine.

The present invention relates to the use of an alkali-stable protein, wherein at least one asparagine residue has been mutated to an amino acid other than glutamine or aspartic acid, which mutation confers an increased chemical stability at pH-values of up to about 13-14 compared to the parental molecule. The protein can for example be derived from a protein capable of binding to other regions of the immunoglobulin molecule than the complementarity determining regions (CDR), such as protein A, and preferably the B-domain of Staphylococcal protein A. The invention also relates to a matrix for affinity separation, which comprises an immunoglobulin-binding protein as ligand coupled to a solid support, in which protein ligand at least one asparagine residue has been mutated to an amino acid other than glutamine.

Provided are Staphylococcal protein A (SpA) variants for binding immunoglobulin (Ig), comprising a polypeptide which varies by one or more amino acids from the amino acid sequence of a natural variable heavy chain III (“VH3”) Ig-Fab binding region (“binding region”) of SpA, wherein the polypeptide exhibits a different binding specificity for Ig-Fab than does SpA or exhibits a different binding specificity for a non-Ig target molecule than does SpA. Further provided are methods of making the variants and methods of using the variants, as well as native SpA, for in purification of Ig as well as diagnostic and therapeutic intervention.

A polypeptide is provided, which has a binding affinity for HER2 and which is related to a domain of staphylococcal protein A (SPA) in that the sequence of the polypeptide corresponds to the sequence of the SPA domain having from 1 to about 20 substitution mutations. Nucleic acid encoding the polypeptide, as well as expression vector and host cell for expressing the nucleic acid, are also provided. Also provided is the use of such a polypeptide as a medicament, and as a targeting agent for directing substances conjugated thereto to cells overexpressing HER2. Methods, and kits for performing the methods, are also provided, which methods and kits rely on the binding of the polypeptide to HER2.

Embodiments concern methods and compositions for treating or preventing a bacterial infection, particularly infection by a Staphylococcus bacterium. Aspects include methods and compositions for providing a passive immune response against the bacteria. In certain embodiments, the methods and compositions involve an antibody that binds Staphylococcal protein A (SpA).

A polypeptide is provided, which has a binding affinity for HER2 and which is related to a domain of staphylococcal protein A (SPA) in that the sequence of the polypeptide corresponds to the sequence of the SPA domain having from 1 to about 20 substitution mutations. Nucleic acid encoding the polypeptide, as well as expression vector and host cell for expressing the nucleic acid, are also provided. Also provided is the use of such a polypeptide as a medicament, and as a targeting agent for directing substances conjugated thereto to cells overexpressing HER2. Methods, and kits for performing the methods, are also provided, which methods and kits rely on the binding of the polypeptide to HER2.

Provided are Staphylococcal protein A (SpA) variants for binding immunoglobulin (Ig), comprising a polypeptide which varies by one or more amino acids from the amino acid sequence of a natural variable heavy chain III (“VH3”) Ig-Fab binding region (“binding region”) of SpA, wherein the polypeptide exhibits a different binding specificity for Ig-Fab than does SpA or exhibits a different binding specificity for a non-Ig target molecule than does SpA. Further provided are methods of making the variants and methods of using the variants for in purification of Ig as well as diagnostic and therapeutic intervention.

The invention relates to a preparation method and application of an electrochemical immunosensor for a biomarker, namely a recombinant proprotein convertase subtilisin kexin type 9 (pcsk9) protein for predicting and diagnosing cardiovascular diseases, and belongs to the technical field of electrochemical detection. The preparation method is characterized by comprising the following steps: firstly, performing nitrogen doping on a graphene nanobelt (n-gnrs), performing amination on a fullerene-palladium-platinum nanoparticle (n-C60 / pdpt) composite material and a staphylococcus aureus a protein (spa), and performing layer-by-layer self-assembling so as to immobilize a pcsk9 antibody (ab1); mixing the synthesized nanoparticle-poly-methylene blue (pt-pmb) with the pcsk9 antibody (ab2), and preparing a nano beacon, thereby obtaining the electrochemical immunosensor for pcsk9 protein detection. The electrochemical immunosensor has the advantages of being high in sensitivity, good in specificity and rapid and convenient in detection. A novel method for pcsk9 protein detection is provided, and useful information is provided for clinical prediction and diagnosis on cardiovascular diseases.

The present invention relates to a novel fusion protein comprising Staphylococcal protein A and mussel adhesive protein, a biochip comprising a solid substrate to which the fusion protein is attached, and a method for detecting a target antigen in a biological sample using the biochip. Furthermore, the present invention relates to a polynucleotide encoding the fusion protein, a recombinant vector comprising the polynucleotide, a transformed cell comprising the recombinant vector, and a method of preparing the fusion protein by transformed cell comprising the recombinant vector.

The invention provides a protein A immunoadsorption material which eliminates pathogenic antibody and the compound from plasma and the a preparation method. The invention discloses a polymer material coupled by agarose gel and staphylococcal protein A (SPA) and the material takes the agarose gel as vector matrix and takes polyamines reagent as a space arm, then reacting with glutaraldehyde and finally coupling with the protein A and blocking, reducing a double bond after being activated by epoxy bromopropane. The product has the high safety; by making use of the produce, the pathogenic antibody, more particularly, the immune compound pathogenic antibody can be effectively and selectively eliminated from the plasma of patients; and the invention can be applied to clinical immunoadsorption treatments.

A polypeptide is provided, which has a binding affinity for HER2 and which is related to a domain of staphylococcal protein A (SPA) in that the sequence of the polypeptide corresponds to the sequence of the SPA domain having from 1 to about 20 substitution mutations. Nucleic acid encoding the polypeptide, as well as expression vector and host cell for expressing the nucleic acid, are also provided. Also provided is the use of such a polypeptide as a medicament, and as a targeting agent for directing substances conjugated thereto to cells overexpressing HER2. Methods, and kits for performing the methods, are also provided, which methods and kits rely on the binding of the polypeptide to HER2.

The present invention relates to preparation of protein, especially the immune detection and amplifying system for preparing coupling agent and its preparation and application. The present invention is one kind of immune signal amplifying system including the compound of staphylococcal protein A (SPA) and streptomycin antibiotic. The system has high detection sensitivity, short operation time.

A chemical luminous ligand analysis method for quantitatively checking human antibodies belongs to the biomedical technical field, which comprises: using the generality that staphylococcal protein A (SPA) can react with Fc point of IgG in mice; using the monoclonal antibody of high affinity and specificity to prepare a standard product to establish a standard curve; respectively reacting the monoclonal antibody and the antibody in the sample with enzyme-labeled antigens; using the nanometer magnetic particles coated by SPA as ligand to separate solid and liquid; and using enzymatic chemical illumination reaction catalyzed by horseradish peroxidase (HRP) to process illumination check; thereby establishing a chemical luminous ligand quantitative analysis on human antibodies. The chemical luminous ligand analysis method is suitable for quantitatively checking all human antibodies, for resolving the problem of prior art while most human antibodies can not be accurately and quantitatively checked, avoiding cross reaction and avoiding false negative result or false positive result.

The invention discloses a hydrogel material for nervus centralis repair and a method for making the same. The method comprises the following steps of: mixing polyethylene glycol water solution with density of 1 to 5 weight percent with polylysine or staphylococcal protein A and nanometer ferric oxide water solution with the density of 0.1 to 2 weight percent, oscillating and stirring; adding glutaral pentanedial solution until reaching final density of 1 to 4 weight percent of the glutaral pentanedial, and reacting for 2 to 24 hours below 4 DEG C; freezing and drying; and cleaning as well as renewly freezing and drying. The hydrogel material for nervus centralis repair made in the method has the advantages of good biocompatibility, rheological characteristics and mechanical properties similar to the nervus centralis tissue, no secondary damage to tissue after implantation of the nervus centralis, and is capable of regulating biodegradability in a large scale and is applicable to nervus repair. And the making method has simple process, low cost, and is suitable for large-scale production.

The invention relates to a staphylococcal protein A (SPA) immunoadsorption material used for blood purification and a preparation method thereof. The invention discloses a macromolecule material which is coupled by agarose gel and SPA. The material is prepared by taking the agarose gel as carrier substrate which is reacted with epoxy bromopropane to obtain the epoxy-based active carrier; after that, polyamines reagent is taken as a space arm which is then reacted with carbonyl diimidazole and is then coupled with the SPA. The material has the advantages of short synthesis time, safe preparation, strong specificity of product, high adsorption efficiency and good regeneration performance to immunoglobulin and the compound thereof, and being able to be applied to clinic immunoadsorption cure.

The invention relates to colloidal gold test paper for rapidly detecting an antibody of a porcine reproductive and respiratory syndrome virus (PRRSV). The test paper is characterized in that: the test paper is composed of a sample pad, a combination pad, a nitrocellulose membrane and an absorption pad which are sequentially attached to a nonabsorbent supporting sheet, wherein the combination pad is covered by PRRSVNsp9 proteins marked by colloidal gold; and the nitrocellulose membrane is respectively covered by a detection line T line composed of staphylococcus proteins A (SPA) and a quality control line C line composed of PRRSVNsp9 monoclonal antibodies 2D6. According to the invention, by utilizing an immunological principle that antigens and antibodies can be specially combined, the antigen marked by the colloidal gold is combined with the corresponding antibody and the conjugate of the antigen and the antibody is combined with the SPA to form an antigen-antibody-SPA conjugate, so as to form a color reaction on the nitrocellulose membrane (NC membrane) to be observed by naked eyes. The test paper provided by the invention has the characteristics of simplicity, fastness, sensitivity, good specificity and the like. And the price is low, so that the test paper is applicable to basically or clinically detecting the antibody of the porcine reproductive and respiratory syndrome virus. The test paper has the obvious advantages of strong specificity, high flexibility, simplicity in operation and fastness in diagnosis, and can be used for rapidly diagnosing the porcine reproductive and respiratory syndrome virus.

The invention relates to a method of isolating an immunoglobulin, comprising the steps of:a) providing a separation matrix comprising multimers of immunoglobulin-binding alkali-stabilized Protein A domains covalently coupled to a porous support,b) contacting a liquid sample comprising an immunoglobulin with the separation matrix,c) washing said separation matrix with a washing liquid,d) eluting the immunoglobulin from the separation matrix with an elution liquid, ande) cleaning the separation matrix with a cleaning liquid,wherein the alkali-stabilized Protein A domains comprise mutants of a parental Fc-binding domain of Staphylococcus Protein A (SpA), as defined by, or having at least 80% such as at least 90%, 95% or 98% identity to, SEQ ID NO 51 or SEQ ID NO 52, wherein the amino acid residues at positions 13 and 44 of SEQ ID NO 51 or 52 are asparagines and wherein at least the asparagine residue at position 3 of SEQ ID NO 51 or 52 has been mutated to an amino acid selected from the group consisting of glutamic acid, lysine, tyrosine, threonine, phenylalanine, leucine, isoleucine, tryptophan, methionine, valine, alanine, histidine and arginine.

The invention discloses an immunoglobulin-binding protein comprising one or more mutated immunoglobulin-binding domains (monomers) of staphylococcal Protein A (E, D, A, B, C) or protein Z or a functional variant thereof, wherein in at least one of the one or more mutated monomers, the asparagine or histidine at the position corresponding to H18 of the B domain of Protein A or of Protein Z has been deleted or substituted with a first amino acid residue which is not proline or asparagine and wherein, if the amino acid residue at position 57 is proline and the amino acid residue at position 28 is asparagine, then the amino acid residue at the position corresponding to H18 of the B domain of protein A or of protein Z is not serine, threonine or lysine.

The present invention discloses a performance improved recombination staphylococcus aureus protein A affinity ligand and a construction method thereof. The present invention adopts a molecular biology method. A sequence B of a nature protein A is selected for molecular transformation. A C-terminal of the sequence B is added with two cysteines, so that the protein A can pass through double-locus coupled chromatography matrix to stabilize the connection. Six glycines are added to the end of a second Loop of the sequence B to increase the length and reduce the binding force with an antibody, so that elution conditions are mild. On this basis, resistance performance to high concentration base of the protein A is transformed. Asparagines and phenylalanine at 23rd and 30th positions of the sequence B are respectively replaced by threonine and alanine to obtain a sequence Z with higher alkaline resistance properties. Then, isocaudarner is used for connecting sequence Zs of different numbers head-to-tail in series. The efficient expression system of e. coli is used for overexpression. The expressed recombination protein A is coupled to agarose matrix preparation affinity chromatography fillers and is used for purifying antibodies. Results show that the recombination protein A affinity ligand prepared by the present invention is good in elution performance and alkali resistance.

The present invention concerns methods and compositions for treating or preventing a bacterial infection, particularly infection by a Staphylococcus bacterium. The invention provides methods and compositions for providing a passive immune response against the bacteria. In certain embodiments, the methods and compositions involve an antibody, such as a recombinant antibody, that binds IsdA and / or IsdB polypeptides.

The invention relates to the biological and pharmaceutical industry, and discloses a magnetic bead method for quickly purifying an antibody. The method is characterized in that by using an affinity chromatography principle, carboxyl magnetic beads are activated, and then connected with stphylococcl protein A (SPA) capable of being combined with Fc (fragment crystalline) of IgG (intravenous gamma globulin) molecules of human and various mammals; and obtained Protein A magnetic beads can be used for purifying the antibody in serum, ascites or a cell culture fluid. The Protein A magnetic beads are high in specificity, quick and convenient in the aspect of antibody purification, and can be widely applied to the biological and pharmaceutical industry.

The present invention relates to a classical swine fever antibody PPA-ELISA detection kit and a preparation method thereof. The kit preparation comprises: adopting RT-PCR to amplify a 550 bp gene fragment of E2 gene, wherein the fragment contains 4 neutralizing antigen regions of A, B, C and D; cloning the amplified 550 bp gene fragment into a pMD18-T vector; carrying out identification, wherein the identification result is correct; further directionally cloning the fragment into a PGEX-6P-1 prokaryotic expression vector; transforming BL21 expression bacterial; carrying out IPTG induced expression to obtain recombinant protein expressed in a form of inclusion body; adopting an affinity chromatography method to separate and purify the recombinant protein; adopting immunoblotting to detect antigenicity and specificity of the purified protein; and adopting the recombinant protein as coating antigen, adopting horseradish peroxidase-labeled staphylococcal protein A (SPA) as second antibody, optimizing reaction conditions of indirect ELISA, establishing an indirect ELISA detection method for detection of classical swine fever virus antibody, and assembling the classical swine fever antibody PPA-ELISA detection kit.

The invention discloses a large-scale preparation method of a recombinant staphylococcus aureus vaccine. The method comprises the following steps: opening a protein working seed lot, inoculating the protein working seed lot strain in a conical flask, inoculating first-generation production strain in a seeding tank, performing continuous second-generation cultivation, inoculating the second-generation production strain in a fermentation tank, and performing continuous third-generation cultivation; centrifugally removing supernate from a bacterial solution through a continuous flow after fermentation is completed; and performing bacterium body dissolving and bacterium disruption; performing combination, washing the mixture after combination is completed, performing digestion, purifying the protein digestion solution by adopting a chromatographic column and performing desalination, endotoxin removing, sterile filtration and storage. The preparation method is simple and can be used for preparing recombinant staphylococcus aureus vaccine in a large scale.

The invention discloses a surface-enhanced Raman gold-labeled test paper strip for quickly detecting trace fipronil and a preparation method thereof. A bottom layer of the test paper strip is a support layer, a middle layer of the test paper strip is an adsorption layer, a protection layer is fixed on the adsorption layer, and the adsorption layer sequentially comprises an absorption fibrous layer, a surface-enhanced Raman colloidal gold-labeled antibody fibrous layer, a cellulose membrane layer and a water absorption material layer of a handle end from a test end, wherein the cellulose membrane is coated with fipronil-coupled carrier protein to serve as a detection line and is also provided with a quality control line printed by goat or rabbit anti-mouse IgG (Immunoglobulin G), goat anti-rabbit IgG antibody or SPA (Staphylococcal Protein A). The test paper strip disclosed by the invention is prepared on the basis of a competitive mode, and target objects on a sample and the detectionline are competitively combined with the surface-enhanced Raman colloidal gold-labeled antibody. Tests show that the test paper has stronger specificity and higher sensitivity, the visual detection limit is an ng level, the quantitative detection limit of a Raman instrument is a pg level, and the test paper has no cross reaction with other pesticides or veterinary drugs.

The invention discloses an affinity chromatography medium, and a preparation method and an application thereof. The affinity chromatography medium provided by the invention comprises a solid phase carrier and recombinant staphylococcal protein A coupled with the solid phase carrier. The amino acid sequence of the recombinant staphylococcal protein A is represented by SEQ ID NO:1. The affinity chromatography medium provided by the invention has the advantages of high protein binding specificity, high affinity and high purification efficiency. The affinity chromatography medium can be used for separation and purification of protein.

This invention discloses to a kind of immunofluorescence mark reagent box. It is a reagent box applied for the aspects of immunity mark which uses the green fluorescence protein (GFP) as the indicator, the staphylococcus protein A (SPA) as the antibody. It also discloses the preparation method of the reagent. This invention can applied wide range in the field such as the immunity fluorescence mark, immunity histochemistry, biochemistry analysis, antibody test, protein chip and so on. It can replace the exist product efficiently, its operation is more simple, stability, delicacy and reliability, it is especially the same with the high flux analysis which analyze by the fluorescence analysis instrument.

The invention relates to a colloidal gold testing paper card used for detecting rabies virus antibodies of dogs and cats. By simultaneously coating a rabies virus-associated antigen strip and a staphylococcal protein A (SPA) antibody strip on a nitrocellulose membrane, and labeling the SPA by adopting colloidal gold, a universal testing paper card is obtained. The rabies virus antibodies in serums of the dogs and the cats are detected by using an immunochromatography principle so as to fulfill an aim of detecting the rabies virus antibodies of the dogs and the cats. The testing paper card overcomes the shortages of complicated and cumbersome operation, long testing time, requirement on instruments and incapability of detecting the antibodies in different animals by using the same testing paper card in the prior detecting art.