# US 2010/0167340 A1 · Method to produce a receptor chip using biotinylated protein > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 2010/0167340 A1 - Document ID: US20100167340A1 - Status: Published application - Kind: A1 - Application number: 12/636,434 - Filed: 2009-12-11 - Published: 2010-07-01 - Earliest claimed: 2002-09-02 - Assignees: National Food Research Institute - Inventors: Sachiko Machida; Kiyoshi Hayashi; Ken Tokuyasu; Yoshikiyo Sakakibara; Shigeru Matsunaga - Web page: https://patentgrep.com/patent/US20100167340A1 ## CPC - G01N 33/54353: {with ligand attached to the carrier via a chemical coupling agent (coatings G01N33/54393)} - G01N 33/566: using specific carrier or receptor proteins as ligand binding reagents {where possible specific carrier or receptor proteins are classified with their target compounds} - B01J 2219/00605: the compounds being directly bound or immobilised to solid supports - B01J 2219/00612: the surface being inorganic - B01J 2219/00617: by chemical means - B01J 2219/0063: Other, e.g. van der Waals forces, hydrogen bonding - B01J 2219/00637: by coating it with another layer - B01J 2219/00659: Two-dimensional arrays - B01J 2219/00725: Peptides - C40B 40/10: Libraries containing peptides or polypeptides, or derivatives thereof - G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q) - B01J: CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS - C40B: COMBINATORIAL CHEMISTRY; LIBRARIES, e.g. CHEMICAL LIBRARIES (in silico combinatorial libraries of nucleic acids, proteins or peptides G16B35/00; in silico combinatorial chemistry G16C20/60) ## Abstract 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. ## Claims ### Claim 1 A receptor chip, on which a recombinantly expressed biotinylated receptor protein is immobilized via a factor capable of specifically binding to biotin. ### Claim 2 (depends on 1) A receptor chip according to claim 1, wherein the biotinylated receptor protein is expressed in a bacterial host. ### Claim 3 (depends on 1) A receptor chip according to claim 1, wherein the biotinylated receptor protein is expressed in vitro. ### Claim 4 (depends on 2) A receptor chip according to claim 2, wherein the biotinylation of the receptor protein is carried out within a bacterial host. ### Claim 5 (depends on 2) A receptor chip according to claim 2, wherein the biotinylation of the receptor protein is carried out in vitro after expression of the protein. ### Claim 6 (depends on 4) A receptor chip according to claim 4, wherein the immobilized biotinylated receptor protein is obtained by refolding a biotinylated receptor protein expressed as an inclusion body within a bacterium. ### Claim 7 (depends on 6) A receptor chip according to claim 6, wherein the refolding is carried out in a solution containing a cyclic carbohydrate cycloamylose and a polyoxyethylene detergent. ### Claim 8 (depends on 7) A receptor chip according to claim 7, wherein the degree of polymerization of the cyclic carbohydrate cycloamylose is 17 to 50 or 40 to 150. ### Claim 9 (depends on 8) A receptor chip according to claim 8, wherein the degree of polymerization of the cyclic carbohydrate cycloamylose is 40 to 150. ### Claim 10 (depends on 7) A receptor chip according to claim 7, wherein the polyoxyethylene detergent is polyoxyethylenesorbitan ester, polyoxyethylenedodecyl ether, polyoxyethylenehepta methylhexyl ether, polyoxyethyleneisooctylphenyl ether, polyoxyethylenenonylphenyl ether, polyoxyethylene fatty acid ester, or sucrose fatty acid ester. ### Claim 11 (depends on 6) A receptor chip according to claim 6, wherein the refolding is carried out in a solution containing a cyclic carbohydrate cycloamylose and an ionic detergent. ### Claim 12 (depends on 11) A receptor chip according to claim 11, wherein the degree of polymerization of a cyclic carbohydrate cycloamylose is 17 to 50 or 40 to 150. ### Claim 13 (depends on 12) A receptor chip according to claim 12, wherein the degree of polymerization of a cyclic carbohydrate cycloamylose is 40 to 150. ### Claim 14 (depends on 11) A receptor chip according to claim 11, wherein the ionic detergent is cetyltrimethyl ammonium bromide, dodecyl sodium sulfate, sodium deoxycholate, 3-[(3-cholamidopropyl)dimethylammonio]-1-propane sulfonate, hexadecyltrimethyl ammonium bromide, or myristyl sulfo betaine. ### Claim 15 (depends on 1) A receptor chip according to claim 1, wherein the receptor is selected from the group consisting of scavenger receptors, receptors of the insulin receptor family, receptors of the EGF receptor family, receptors of the PDGF receptor family, receptors of the VEGF receptor family, receptors of the FGF receptor family, growth factor receptors of the NGF receptor family, TGF-(3 super family receptors, Toll-like receptor family, LDL receptor related protein family, and receptors of the G protein coupled receptor family. ### Claim 16 (depends on 15) A receptor chip according to claim 15, wherein the receptor is a scavenger receptor LOX-1. ### Claim 17 (depends on 1) A receptor chip according to claim 1, adapted for detection using surface plasmon resonance, quartz-crystal microbalance, or mass spectrometer. ### Claim 18 A method for producing a receptor chip, comprising the steps of: a) recombinantly expressing a biotinylated receptor protein as an inclusion body within a bacterial host; b) refolding the inclusion body in a solution containing a cyclic carbohydrate cycloamylose and a polyoxyethylene detergent to prepare a soluble biotinylated receptor protein; and c) immobilizing the refolded soluble biotinylated receptor protein to a solid phase via a factor capable of specifically binding to biotin. ### Claim 19 (depends on 18) A method according to claim 18, wherein the degree of polymerization of the cyclic carbohydrate cycloamylose is 17 to 50 or 40 to 150. ### Claim 20 (depends on 19) A method according to claim 19, wherein the degree of polymerization of the cyclic carbohydrate cycloamylose is 40 to 150. ### Claim 21 (depends on 18) A method according to claim 18, wherein the polyoxyethylene detergent is polyoxyethylenesorbitan ester, polyoxyethylenedodecyl ether, polyoxyethyleneheptamethylhexyl ether, polyoxyethyleneisooctylphenyl ether, polyoxyethylenenonylphenyl ether, polyoxyethylene fatty acid ester, or sucrose fatty acid ester. ### Claim 22 (depends on 18) A method according to claim 18, wherein the receptor is selected from the group consisting of scavenger receptors, receptors of the insulin receptor family, receptors of the EGF receptor family, receptors of the PDGF receptor family, receptors of the VEGF receptor family, receptors of the FGF receptor family, growth factor receptors of the NGF receptor family, TGF-β super family receptors, Toll-like receptor family, LDL receptor related protein family, and receptors of the G protein coupled receptor family. ### Claim 23 (depends on 22) A method according to claim 22, wherein the receptor is a scavenger receptor LOX-1. ### Claim 24 (depends on 18) A method according to claim 18, wherein the solid phase is adapted for detection using surface plasmon resonance, quartz-crystal microbalance, or mass spectrometer. ### Claim 25 A method for producing a receptor chip, comprising the steps of: a) recombinantly expressing a biotinylated receptor protein as an inclusion body within a bacterial host; b) refolding the inclusion body in a solution containing a cyclic carbohydrate cycloamylose and an ionic detergent to prepare a soluble biotinylated receptor protein; and c) immobilizing the refolded soluble biotinylated receptor protein to a solid phase via a factor capable of specifically binding to biotin. ### Claim 26 (depends on 25) A method according to claim 25, wherein the degree of polymerization of the cyclic carbohydrate cycloamylose is 17 to 50 or 40 to 150. ### Claim 27 (depends on 26) A method according to claim 26, wherein the degree of polymerization of the cyclic carbohydrate cycloamylose is 40 to 150. ### Claim 28 (depends on 25) A method according to claim 25, wherein the ionic detergent is cetyltrimethyl ammonium bromide, dodecyl sodium sulfate, sodium deoxycholate, 3-[(3-cholamidopropyl)dimethylammonio]-1-propane sulfonate, hexadecyltrimethyl ammonium bromide, or myristyl sulfo betaine. ### Claim 29 (depends on 25) A method according to claim 25, wherein the receptor is selected from the group consisting of scavenger receptors, receptors of the insulin receptor family, receptors of the EGF receptor family, receptors of the PDGF receptor family, receptors of the VEGF receptor family, receptors of the FGF receptor family, growth factor receptors of the NGF receptor family, TGF-β super family receptors, Toll-like receptor family, LDL receptors related protein family, and receptors of the G protein coupled receptor family. ### Claim 30 (depends on 29) A method according to claim 29, wherein the receptor is a scavenger receptor LOX-1. ### Claim 31 (depends on 25) A method according to claim 25, wherein the solid phase is adapted for detection using surface plasmon resonance, quartz-crystal microbalance, or mass spectrometer. ### Claim 32 (depends on 18, 25) A receptor chip produced by a method according to claim 18 or 25. ### Claim 33 (depends on 16) A method for detecting modified LDL, an abnormal cell, or a bacterium, using a receptor chip according to claim 16. ### Claim 34 (depends on 23, 30) A receptor chip produced by a method according to claim 23 or 30. ### Claim 35 (depends on 34) A method for detecting modified LDL, an abnormal cell, or a bacterium, using a receptor chip according to claim 34. ### Claim 36 (depends on 18, 25) A detection kit, comprising a receptor chip produced by a method according to claim 18 or 25. ### Claim 37 (depends on 16) A detection kit, comprising a receptor chip according to claim 16. ### Claim 38 (depends on 23, 30) A detection kit, comprising a receptor chip produced by a method according to claim 23 or 30. ### Claim 39 (depends on 34) A detection kit, comprising a receptor chip according to claim 34.