33 results about "Glycoproteomics" patented technology

A diagnostic method for determining the absence or presence of a disease is provided. The method includes assaying the amount and/or types of glycopeptides in a sample from a subject, and comparing these to the amount and types of reference glycopeptides. The method may include the use of a stable isotope label, affinity selection, immunoaffinity chromatography, and glycoproteomics techniques, to identify and quantify changes in glycosylated peptides or glycosylated proteins associated with cancers such as malignant lymphoma or breast cancer, to monitor patient's response to therapy, and to monitor disease recurrence.

The invention relates to a bonding polysaccharide type hydrophilic chromatographic stationary phase as well as a preparation method and an application thereof. A structure of the bonding polysaccharide type hydrophilic chromatographic stationary phase is shown in the specification, wherein SiO2 is silica gel, saccharide is polysaccharide with molecular weight ranging from hundreds to tens of thousands. The invention provides the preparation method of the stationary phase material, wherein the preparation method comprises the following step: modifying the surface of the silica gel and bonding with the polysaccharide in an aqueous liquor system by taking carbonyl dimidazole (CDI) as a connecting arm to prepare the bonding polysaccharide type chromatographic stationary phase. Compared with the conventional bonding saccharide type stationary phase preparation method, the stationary phase preparation method disclosed by the invention is more efficient, convenient and environment-friendly. And meanwhile, the bonding polysaccharide type hydrophilic chromatographic stationary phase has good separating selectivity, repeatability and stability, has great application potentiality in separating polar compounds of other types, and is widely applied in saccharide separating and saccharide-modifying proteomics.

The invention belongs to the field of the glycoproteomics and glycomics, and relates to a method for rapid enzymolysis release, solid phase concentration and mass spectrometry of N-glycan. The methodcomprises the following steps of absorbing a solution containing a glycoprotein sample by using a degreasing cotton material, and performing high-temperature rapid PNGase F enzymolysis on the cotton to release N-glycan; adjusting solution environment after the enzymolysis is finished, and adsorbing free glycan on the degreasing cotton material by utilizing the high-density hydroxy and hydrogen bond of the glycan and like effects on the surface of the degreasing cotton material, and then cleaning and removing non-glycan molecules such as protein, polypeptide and like which are not bonded with the degreasing cotton material, and finally eluting the adsorbed glycan with the aqueous solution, and feeding into mass spectrometry. Through the method disclosed by the invention, the used material is cheap and easy to obtain, the preparation is easy, the operation is simple and convenient, the high-speed enzymolysis of the N-glycoprotein and the high-selective concentration and high-sensitive mass spectrometry of the N-glycan can be realized.

The invention specifically relates to preparation of a microporous polymer coated hydrophilic resin and application thereof in glycopeptide enrichment. The preparation method includes: firstly preparing porous epoxy resin microspheres that have a particle size of 8-15microm and contain a plurality of epoxy functional groups on the surfaces, then adding m-phenylenediamine and isophthalaldehyde, andtaking acetic acid as the catalyst to prepare the hydrophilic resin coated with a layer of microporous organic polymer on the surface. The preparation conditions of the hydrophilic resin are simple and the reaction is mild. And finally, a standard glycoprotein (IgG) enzymolysis liquid is adopted as the sample to investigate the enrichment performance of hydrophilic resin to glycopeptide, and thehydrophilic resin can be further applied to glycoproteomics analysis in rat liver.

The invention belongs to the technical field of glycopeptide/glycoprotein enrichment materials and analysis, and relates to a sugar chain releasable glycosylated peptide fragment/protein enrichment material based on a hydrazide strategy, a preparation method of the material and an application of the material. The surface of a matrix micro-sphere is bonded with a hydrazide compound containing disulfide bonds to form a functional material with a hydrazide group on the surface, wherein the functional material can be used for glycopeptide enrichment. The disulfide bonds are led to the surface of the material to realize sugar chain releasable enrichment of glycopeptide/glycoprotein, sugar chains are in covalent binding, and the material has the advantage of high enrichment efficiency and overcomes the shortcoming that the sugar chains are not easily released in a traditional hydrazide method and cannot be used for O-glycosylated peptide fragment/protein enrichment. The sugar chain releasable enrichment material is easy to prepare and good in enrichment selectivity, is widely applicable to N-glycosylated and O-glycosylated peptide fragment/protein enrichment and has a high practical value in the fields of glycoproteomics and the like.

The invention discloses an MALDI-MS based and stable isotope labeled N-glycan rapid quantitative analysis method and application of the MALDI-MS based and stable isotope labeled N-glycan rapid quantitative analysis method, and belongs to the field of glycoproteomics analysis. The method comprises the following steps that (1) under the assistance of microwave, PNGase F makes N-glycans released in a form of osamine, (2) d0/d5-benzoyl chloride has a nucleophilic reaction with osamine respectively and then purified, (3)d0-benzoyl chloride and d5-benzoyl chloride-derived glycans are mixed at a molar ratio of 1: 1, (4) mixed d0/d5-benzoyl chloride derived N-glycans are subjected to a methylamine reaction, (5) MALDI-MS detection is performed, (6) data are analyzed; the MALDI-MS based and stable isotope labeled N-glycan rapid quantitative analysis method and the application of the MALDI-MS based and stable isotope labeled N-glycan rapid quantitative analysis method disclosed by the invention realize simultaneous quantitative analysis of neutral sugar and acid sugar, reduce the experimental cost, improve the experimental efficiency and shorten the reaction time, and have a wider linear quantitative range, can be used for serum glycan analysis, and have the medical application values.

Methods for producing isotopically-labelled peptides are provided. Aspects of the method include: contacting a sample including a metabolically tagged protein with a cleavable probe to produce a probe-protein conjugate; separating the probe-protein conjugate from the sample; digesting the probe-protein conjugate to produce a probe-peptide conjugate; and cleaving a cleavable linker to release an isotopically labelled peptide. The method may further include: identifying a predetermined isotopic pattern in a mass spectrum; determining an amino acid sequence of the isotopically labelled peptide; and identifying the site of protein glycosylation based on the determined amino acid sequence. Also provided are cleavable probes for practicing the subject methods, described by the Formula: A-L-(M-Z) where A is an affinity tag, L is a cleavable linker, M is an isotopic label and Z is a chemoselective tag capable of cross-linking a metabolically tagged protein. Compositions and kits for practicing the subject methods are also provided.

The invention more specifically relates to a method for preparing a hydrophilic resin through a multilayer self-assembly method. The hydrophilic resin can be used for enriching glycopeptides. A porousepoxy resin with the particle size in a range of 100-500 [mu]m is prepared through a suspension polymerization method and is taken as a resin adsorbent, and then, multilayer polysaccharide (hyaluronic acid and chitosan) coats the surface of the porous epoxy resin through a multilayer self-assembly method to prepare the hydrophilic resin. A standard glycoprotein enzymatic hydrolysate is selected as a sample to study the enrichment performance of the hydrophilic resin to glycopeptides. The hydrophilic resin is further applied to glycoproteomics analysis of protein that is extracted from the ratliver.

The invention relates to a polymer chain-modified silica gel matrix hydrophilic interaction chromatography stationary phase as well as a preparation method and application thereof. Silica gel particles are taken as a matrix, and a hydroxyl-containing allyl monomer and methylene bisacrylamide are grafted on the surface of silica gel by means of a click copolymerization reaction of thiol-ene, so that a chromatography stationary phase which has a three-dimensional dendritic polymer chain, is rich in hydroxyl and amide hydrophilic functional groups and has hydrophilic interaction can be formed. The hydroxyl and amide hydrophilic functional groups and the like are introduced according to the method provided by the invention, so that not only are the defects that the traditional post-modification method is complicated in steps and low in reaction efficiency overcome, but a rich hydrophilic layer can be also formed by the three-dimensional polymer chain-shaped structure, therefore, the hydrophilic chromatography stationary phase has better advantages. The liquid chromatography stationary phase provided by the invention is novel in structure and excellent in hydrophilic performance, can be widely used for separation of various samples and selective enrichment and separation of glycopeptides, and has a very high practical value in the fields of separation analysis, glycoproteomics, and the like.

The invention relates to a hydrophilic macroporous chitosan integral material prepared by using a freeze drying method and application thereof. According to the specific preparation process of the material, chitosan is used as a monomer, polyethylene glycol diglycidyl ether is used as a cross-linking agent, an acetic acid solution is used as a solvent, and the hydrophilic chitosan integral material is prepared through the freeze-drying method. The integral material disclosed by the invention is low in cost, green and environment-friendly, has a good specific adsorption effect on glycopeptide in a complex sample, and shows good application potential in glycoproteomics research.

The invention relates to a preparation method of a glycoprotein microreactor for boron affinity surface imprinting of a mesoporous molecular sieve. According to the method, an SBA-15 mesoporous molecular sieve is used as a carrier, 2,3-difluoro-4-formylphenylboronic acid is used as a monomer, and N-acetylneuraminic acid is used as a fragment template to prepare the molecular sieve-based surface oriented imprinting polymer (SBA-15-coated MIP). According to the micro-reactor, protein in a sample is extracted by utilizing a size exclusion effect, the protein is subjected to rapid enzymolysis based on a nano confinement effect, and a peptide fragment containing a sugar chain is selectively enriched by utilizing a specific recognition effect of molecular imprinting. According to the method, the used materials are low in price and easy to obtain, preparation is easy, operation is easy and convenient, the three sample pretreatment processes of protein extraction, rapid protein enzymolysis and glycopeptide enrichment can be integrated, and the pretreatment time of the glycoproteomics sample can be effectively shortened.

The invention belongs to the field of glycoproteomics analysis, and relates to a method for complete glycopeptide derivation and charge transfer fragmentation mass spectrometry. According to the method, all carboxyl groups of the N-glycopeptide are subjected to tertiary amine small molecule derivation and then sent to charge transfer fragmentation mass spectrometry for analysis. The method is simple in step, convenient to operate, rapid and efficient, and the high-sensitivity and high-reliability analysis of the complete glycopeptide can be achieved.

The invention provides a glycoproteomics sample pretreatment device and method.The glycoproteomics sample pretreatment device comprises a pipette head, absorbent cotton, HILIC filler, a C18 extraction membrane and mixed ion exchange resin, and the pipette head is sequentially filled with the absorbent cotton, the HILIC filler, the C18 extraction membrane and the mixed ion exchange resin from bottom to top. According to the glycoproteomics sample pretreatment device disclosed by the invention, through the synergistic combination of the degreasing cotton, the HILIC filler, the C18 extraction membrane and the mixed ion exchange resin, the whole process of pre-enrichment, reductive alkylation, enzymolysis, peptide fragment desalination and complete glycopeptide enrichment of a glycoprotein sample can be integrally realized. A sample pretreatment method based on the glycoproteomics sample pretreatment device can effectively reduce sample loss caused in the sample treatment process and shorten the sample treatment time, large-scale glycoproteomic analysis of trace protein samples can be realized, and the analysis sensitivity of complete glycopeptides is improved.