# US 8,785,143 B2 · Method for immobilizing streptavidin on a self-assembled monolayer > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 8,785,143 B2 - Document ID: US8785143B2 - Status: Granted - Kind: B2 - Application number: 13/629,121 - Filed: 2012-09-27 - Published: 2013-01-24 - Granted: 2014-07-22 - Earliest claimed: 2010-08-30 - Assignees: Panasonic Healthcare Co., Ltd. - Applicants: Panasonic Corporation - Inventors: Yukari Hataoka - Web page: https://patentgrep.com/patent/US8785143B2 ## CPC - G01N 33/54353: {with ligand attached to the carrier via a chemical coupling agent (coatings G01N33/54393)} - C07K 17/14: Peptides being immobilised on, or in, an inorganic carrier - A61K 2039/625: {binding through the biotin-streptavidin system or similar} - C07K 2319/20: containing a tag with affinity for a non-protein ligand - G01N 33/53: Immunoassay; Biospecific binding assay; Materials therefor - G01N 33/54306: {Solid-phase reaction mechanisms} - G01N 2610/00: Assays involving self-assembled monolayers [SAMs] - 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) - C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00) - A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D) ## Abstract Provided are a method for increasing an amount of streptavidin to be immobilized on the self-assembled monolayer and a sensor which comprises streptavidin immobilized with the method. The method of the current technology is characterized by that one molecule of an amino acid is interposed between the self-assembled monolayer and the molecule of streptavidin. ## Claims ### Claim 1 A method for immobilizing a streptavidin on a self-assembled monolayer, comprising the following step (a) and step (b) in this order: a step (a) of preparing a substrate comprising one molecule of an amino acid and the self-assembled monolayer, wherein, the one molecule of the amino acid is bound to the self-assembled monolayer through a peptide bond represented by the following chemical formula (I): wherein R represents a side chain of the one molecule of the amino acid, and wherein the one molecule of the amino acid is selected from the group consisting of Lysine, Histidine, Phenylalanine, Cysteine, Glycine, Methionine, Glutamate, Tyrosine, Alanine, Serine, Aspartate, Asparagine, Leucine, and Tryptophan, and a step (b) of supplying the streptavidin to the substrate to form a peptide bond represented by the following chemical formula (II) as a result of reaction between the carboxyl group of the one molecule of the amino acid and the amino group of the streptavidin wherein R represents a side chain of the one molecule of the amino acid. ### Claim 2 (depends on 1) A method according to claim 1, wherein the step (a) comprises the following step (a1) and step (a2): a step (a1) of preparing a substrate comprising a self-assembled monolayer on the surface thereof, the self-assembled monolayer having a carboxylic acid at one end, and a step (a2) of supplying the one molecule of the amino acid to form a peptide bond represented by the chemical formula (I) as a result of reaction between the carboxylic group of the one end of the self-assembled monolayer and the amino group of the one molecule of the amino acid. ### Claim 3 (depends on 1) A method according to claim 1, further comprising the following step (ab) between the step (a) and the step (b): a step (ab) of activating the carboxyl group of the one molecule of the amino acid with a mixture of N-Hydroxysuccinimide and 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride. ### Claim 4 (depends on 2) A method according to claim 2, further comprising the following step (a1a) between the step (a1) and the step (a2): a step (a1a) of activating the carboxyl group of the self-assembled monolayer with a mixture of N-Hydroxysuccinimide and 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride. ### Claim 5 (depends on 1) A method according to claim 1, wherein the chemical formula (II) is represented by the following chemical formula (III) wherein R represents a side chain of the one molecule of the amino acid. ### Claim 6 (depends on 1) A method according to claim 1, wherein the one molecule of the amino acid is selected from the group consisting of Lysine, Histidine, Phenylalanine, Cysteine, Glycine, Methionine, Glutamate, Tyrosine, Alanine, Serine, Aspartate, and Asparagine. ### Claim 7 (depends on 1) A method according to claim 1, wherein the one molecule of the amino acid is selected from the group consisting of Lysine, Histidine, Phenylalanine, Cysteine, Glycine, Methionine, Glutamate, Tyrosine, Alanine, and Serine. ### Claim 8 A method for detecting or quantifying a target substance contained in a sample with a sensor, comprising the following steps (a) to (d) in this order, a step (a) of preparing a sensor substrate comprising one molecule of an amino acid and a self-assembled monolayer, wherein, the one molecule of the amino acid is bound to the self-assembled monolayer through a peptide bond represented by the following chemical formula (I): wherein R represents a side chain of the one molecule of the amino acid, and wherein the one molecule of the amino acid is selected from the group consisting of Lysine, Histidine, Phenylalanine, Cysteine, Glycine, Methionine, Glutamate, Tyrosine, Alanine, Serine, Aspartate, Asparagine, Leucine, and Tryptophan; a step (b) of supplying the streptavidin to the sensor substrate to form a peptide bond represented by the following chemical formula (II) as a result of reaction between the carboxyl group of the one molecule of the amino acid and the amino group of the streptavidin wherein R represents a side chain of the one molecule of the amino acid, a step (c) of supplying the sample to the sensor substrate of formula (II) to cause the target substance to bind to the streptavidin, and a step (d) of detecting or quantifying the target substance. ### Claim 9 (depends on 8) A method according to claim 8, wherein, the step (c) comprises the following steps (c1) and (c2): a step (c1) of binding an antibody to the streptavidin, the antibody modified with biotin, and a step (c2) of binding the target substance to the antibody. ### Claim 10 (depends on 8) A method according to claim 8, wherein the chemical formula (II) is represented by the following chemical formula (III) wherein R represents a side chain of the one molecule of the amino acid. ### Claim 11 (depends on 8) A method according to claim 8, wherein the one molecule of the amino acid is selected from the group consisting of Lysine, Histidine, Phenylalanine, Cysteine, Glycine, Methionine, Glutamate, Tyrosine, Alanine, Serine, Aspartate, and Asparagine. ### Claim 12 (depends on 8) A method according to claim 8, wherein the one molecule of the amino acid is selected from the group consisting of Lysine, Histidine, Phenylalanine, Cysteine, Glycine, Methionine, Glutamate, Tyrosine, Alanine, and Serine.