Sensor for detecting saccharide and manufacturing method thereof and detection method of glycated hemoglobin using the same

• US 9,897,920 B2
• Filed: 03/11/2015
• Issued: 02/20/2018
• Est. Priority Date: 10/21/2014
• Status: Active Grant

• Current Assignee: Ajou University Industry, LG Electronics, Inc. (LG Corporation)
• Original Assignee: Industry-Academic Cooperation Foundation Yonsei University, LG Electronics, Inc. (LG Corporation)
• Inventors: Ku, Yunhee, Kim, Moosub, Yang, Yongju, Song, Seung Yeon, Lee, Kang Sun, Yoon, Hyun Chul, Han, Yong Duk, Park, Yoo Min
• Primary Examiner(s): Visone, Thomas J
• Application Number:
  US14/645,009
• Publication Number:
  US 20160109370A1
• Time in Patent Office: 1,077 Days
• Field of Search: None
• CPC Class Codes: G01N 21/78 producing a change of colour
• CPC Class Codes: G01N 2400/00 Assays, e.g. immunoassays or enzyme assays, involving carbohydrates
• CPC Class Codes: G03F 7/027 Non-macromolecular photopolymerisable compounds having carbon-to-carbon double bonds, e.g. ethylenic compounds G03F7/075 takes precedence
• CPC Class Codes: G03F 7/2002 with visible light or UV light, through an original having an opaque pattern on a transparent support, e.g. film printing, projection printing; by reflection of visible or UV light from an original such as a printed image

Abstract

The present disclosure relates to a sensor for detecting saccharide and manufacturing method thereof and detection method of glycated hemoglobin using the same wherein the sensor for detecting saccharide includes a reactive layer synthesized with hydrogel having a boronic acid-modified 3D mesh structure. A selective and sensitive detection of the glycated protein including glucose or glycated hemoglobin (HbA_1c) in blood through high level of signal may be enabled, and the durability of the sensor for detecting saccharide may be enhanced.

Claims

1. A sensor for detecting a saccharide, the sensor comprising:

a reactive layer comprising a hydrogel having a boronic acid-modified three-dimensional mesh structure; and

a porous membrane substrate fixed with the reactive layer,

wherein the hydrogel comprises 8.00%-10.00% (w/v) of N-(2-hydroxyethyl) acrylamide, 0.40%-0.50% (w/v) of N,N′

-methylene-bisacrylamide, and 0.10%-2.00% (w/v) of 3-(acrylamido) phenylboronic acid.

2. The sensor of claim 1, wherein the saccharide is a saccharide portion of glycated protein or glucose.

3. The sensor of claim 2, wherein the glycated protein is glycated hemoglobin (HbA_1c).

4. The sensor of claim 1, wherein the porous membrane substrate is a substrate using a membrane for a LFA (Lateral Flow Assay) formed of glass fibers.

5. A method for manufacturing the sensor of claim 1, the method comprising:

injecting a polymer monomer, a polymer cross-linker, boronic acid, and a photo-polymerization reaction initiator into the porous membrane substrate;

patterning the porous membrane substrate with a photomask;

irradiating UV (ultraviolet) onto the patterned porous membrane substrate;

washing the UV-irradiated porous membrane substrate;

and drying the washed porous membrane substrate,

wherein the polymer monomer is N-(2-hydroxyethyl)acrylamide, the polymer cross-linker is N,N′

-methylene-bisacrylamide, and the boronic acid is 3-(acrylamido)phenylboronic acid, and wherein the N-(2-hydroxyethyl)acrylamide, the N,N′

-methylene-bisacrylamide, the 3-(acrylamido)phenylboronic acid and the photo-polymerization reaction initiator are mixed in a dimethyl sulfoxide solvent prior to injection at a weight ratio of 8.00-10.00% (w/v) of the N-(2-hydroxyethyl)acrylamide, 0.40-0.50% (w/v) of the N,N′

-methylene-bisacrylamide, 0.10-2.00% (w/v) of the 3-(acrylamido)phenylboronic acid, and 0.05-0.14%(w/v) of the photo-polymerization reaction initiator.

6. The method of claim 5, wherein the porous membrane substrate is a substrate using a membrane for a LFA (Lateral Flow Assay) formed of glass fibers.

7. A method for detecting glycated hemoglobin, the method comprising:

mixing and reacting glycated hemoglobin and horseradish peroxidase on the sensor of claim 1;

washing non-reactive protein after the reacting of the mixed glycated hemoglobin and horseradish peroxidase;

inducing a color development by dripping a chromogen working solution including a chromogen and a substrate with respect to the horseradish peroxidase onto the sensor;

digitizing a degree of the color development; and

outputting the digitized degree corresponding to a concentration of the detected glycated hemoglobin.

8. The method of claim 7, wherein the substrate is hydrogen peroxide, and the chromogen is any one selected from the group consisting of 3,3′

,5,5′

-tetramethylbenzidine, o-phenylenediamine dihydrochloride, 2,2′

-azinobis[3-ethylbenzothiazoline-6-sulfonic acid]-diammonium salt, 3,3′

-diaminobenzidine tetrahydrochloride, SAT-3 (analog of o-tolidine), N-ethyl-N-(2-hydroxy-3-sulfopropyl)-3-methoxyaniline sodium salt dihydrate, N-ethyl-N-(3-sulfopropyl)-3-methoxyaniline sodium salt monohydrate, N-ethyl-N-(3-sulfopropyl)aniline, sodium salt, N-ethyl-N-(2-hydroxy-3-sulfopropyl)-3,5-dimethoxyaniline sodium salt, N-(2-hydroxy-3-sulfopropyl)-3,5-dimethoxyaniline sodium salt), N,N-Bis(4-sulfobutyl)-3,5-dimethylaniline disodium salt, N-ethyl-N-(2-hydroxy-3-sulfopropyl)-3,5-dimethylaniline sodium salt monohydrate, N,N-Bis(4-sulfobutyl)-3-methylaniline disodium salt, N-ethyl-N-(2-hydroxy-3-sulfopropyl)-3-methylaniline sodium salt dihydrate and N-ethyl-N-(3-sulfopropyl)-3-methylaniline sodium salt monohydrate.

9. The method of claim 7, wherein the mixing and reacting the glycated hemoglobin and the horseradish peroxidase comprises:

dropping 1 to 100 μ

l of a solution, which is prepared by mixing and resolving the horseradish peroxidase and the glycated hemoglobin at a volumetric ratio of 1;

1 in a PBS (phosphate-buffered saline) solution to a concentration of 1 to 10 μ

g/ml, onto the sensor; and

reacting the dropped solution for 10 to 600 seconds.

10. The sensor of claim 1, wherein the hydrogel is synthesized by a photopolymerized reaction with a polymer monomer, a polymer cross-linker, and boronic acid in the porous membrane substrate, wherein the polymer monomer is N-(2-hydroxyethyl)acrylamide, the polymer cross-linker is N,N′

-methylene-bisacrylamide, and the boronic acid is 3-(acrylamido) phenylboronic acid.

11. The sensor of claim 10, wherein concentration (w/v) of the polymer monomer and the boronic acid for the synthesis into the reactive layer is within a range of 4:

1 to 100;

1.