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Method and kit for detecting pathogenic microorganism

Foreign code F190009868
File No. AF19-14TW
Posted date Jul 29, 2019
Country Taiwan
Application number 106130237
Gazette No. 201812297
Date of filing Sep 5, 2017
Gazette Date Apr 1, 2018
Priority data
  • P2016-172515 (Sep 5, 2016) JP
  • P2017-099579 (May 19, 2017) JP
Title Method and kit for detecting pathogenic microorganism
Abstract Provided is a method for detecting pathogenic microorganisms in a biological sample, which is a technique that can be used to perform high-sensitivity detection of pathogenic microorganisms, such as influenza virus, etc., the method including: an introducing step for introducing a hydrophilic solvent that contains the biological sample and substances that serve as substrates for reactions involving enzymes that are present on surfaces of the pathogenic microorganisms or in the interiors thereof into a space between a lower-layer section in which a plurality of accommodating sections that can accommodate the pathogenic microorganisms are formed and an upper-layer section that faces a surface in which the accommodating sections are formed in the lower-layer section; an encapsulating step for introducing a hydrophobic solvent into the space and forming, in the accommodating sections, droplets of the hydrophilic solvent that are coated by the hydrophobic solvent enveloping the pathogenic microorganisms and the substances; and a detecting step for optically detecting reaction products that are generated as a result of the reactions between the enzymes and the substances in the droplets, wherein the hydrophilic solvent has a pH value that is greater than the acid dissociation constant (pKa) of the reaction products.
Scope of claims [claim1]
1. A methods, it is examines the self inductance catching an illness original nature microorganism the object or has in the biological sample that the suspicion infection's object separates this pathogenicity microorganism's method, its includes: Inducts the step, it is in may accept the above pathogenicity microorganism a plural number admission section because of the lower level department that having the hydrophobic surface's sidewall each other was separated to form, and in this lower level department forms this admission section facing to the space between upper departments, inducts water affinity solvent the step, but this water affinity solvent will include this biological sample and will become the use to exist in the surface or internal enzyme's this pathogenicity microorganism matrix of response material; Seals into the step, it is inducts the hydrophobic solvent in this space, but in this admission section, forms after the hydrophobic solvent superposition and contains this pathogenicity microorganism and this material the step of bubble of water affinity solvent; The examination step, it is optics examine in this bubble because of this enzyme with the step that of reaction product the response of this material produces; And this water affinity solvent has the acid hydrolysis of this reaction product to leave the constant (pKa) also big pH value.

[claim2]
2. Like request 1 method, this pathogenicity microorganism is the influenza virus, this enzyme for the nerve amine saccharinic acid enzyme, this material is 4- methyl umbrella type bases - α-D - nerve amine saccharinic acid (4-Methylumbelliferyl-N-acetyl-α-D-neuraminic acid), this reaction product is 4- methyl umbrella type colored lactones.

[claim3]
3. 1 kinds of suites, it is uses to examine the self inductance catching an illness original nature microorganism the object or has in the biological sample that the suspicion infection's object separates this pathogenicity microorganism's suite, its includes: The array, its has to be possible to accept this pathogenicity microorganism a plural number admission section, and to this lower level department in forms the surface gap space that because of the lower level department that having the hydrophobic surface's sidewall each other was separated to form this admission section forms, but to approaching upper department; Will become the use to exist in the surface or internal enzyme's this pathogenicity microorganism matrix of response material; Has uses this enzyme to leave the constant (pKa) also big pH value with acid hydrolysis that of the reaction product the response of this material produces the water affinity solvent; And hydrophobic solvent.

[claim4]
4. 1 methods, it is in makes the contact surface contact with the hydrophobic solvent in the water affinity solvent, makes the enzyme, attending to become the material that the matrix that of response uses this enzyme responded, but the method of examination reaction product, this water affinity solvent has the acid hydrolysis of this reaction product to leave the constant (pKa) also big pH value.

[claim5]
5. Like request 4 method, this water affinity solvent includes the pathogenicity microorganism, this enzyme to exist has the matrix to shut off the active enzyme in the surface or the interior this pathogenicity microorganism, this material is the colored matrix, optics examination because of using this enzyme cut-off of the colored matrix the reaction product of production.

[claim6]
6. Like request 5 method, this pathogenicity microorganism is the influenza virus, this enzyme for the nerve amine saccharinic acid enzyme, this colored matrix is 4- methyl umbrella type bases - α-D - nerve amine saccharinic acid (4-Methylumbelliferyl-N-acetyl-α-D-neuraminic acid), this reaction product is 4- methyl umbrella type colored lactones.

[claim7]
7. Like request item 5 or 6 methods, this water affinity solvent includes from infecting this pathogenicity microorganism's object or has the biological sample that the suspicion infection's object separates.
  • Applicant
  • JAPAN SCIENCE AND TECHNOLOGY AGENCY
  • Inventor
  • NOJI HIROYUKI
  • TABATA KAZUHITO
IPC(International Patent Classification)
Reference ( R and D project ) CREST Creation of Nanosystems with Novel Functions through Process Integration AREA
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