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Assays for identifying compounds that modulate bitter taste

By labeling the insoluble carrier particles with a fluorescent dye that complements its absorption wavelength, the problems of insufficient sensitivity and difficulty in visual judgment in the existing technology are solved, and an efficient and simple immunoassay is achieved, which is suitable for POCT.

Active Publication Date: 2016-02-24
DENKA CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Insoluble carrier particles such as gold colloids, platinum colloids, and colored polystyrene particles have the advantage of being visually identifiable. However, even when these insoluble carrier particles are detected with a dedicated device, a significant improvement in detection sensitivity cannot be expected.
On the other hand, when using a ligand labeled with a fluorescent substance or an insoluble carrier particle containing a fluorescent substance, if a dedicated device is used, there is an advantage that a significant improvement in detection sensitivity can be expected, but it cannot be judged visually, even if it is detected The amount of substance is large, and it can be detected in a short time by visual judgment. It also requires the use of a dedicated device, which becomes complicated
[0005] In order to solve this problem, it has been proposed to mix insoluble carrier particles and fluorescent particles that can be judged visually (see Patent Document 1). However, since the ligands on the insoluble carrier particles and the ligands on the fluorescent particles compete with each other, the When the detection substance is very small, the ligand on the fluorescent particle cannot be efficiently combined, so it may not be a high sensitivity
In addition, when considering production efficiency, it is necessary to prepare two types of insoluble carrier particles separately, leaving the problem of low efficiency

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0074] Example 1 Preparation of Visibly Colored Fluorescent Particles Labeled with Fluorescent Pigments Using Colored Polystyrene Particles Having Carboxyl Groups as Functional Groups

[0075] 1. Conjugation of Europium Chelate Fluorescent Labeling Agent

[0076] Colored polystyrene particles (see Table 1 and Figure 1~3 ) was diluted to 0.1%, and ATBTA-Eu as a europium chelate fluorescent labeling agent was added therein 3+ (Tokyo Chemical Industry Co., Ltd.), make ATBTA-Eu 3+ 0.1 mg / mL. After stirring, carbodiimide was added to make it 1%, stirred, and reacted at room temperature for 1 hour. After centrifugation at 7000 g for 30 minutes, the supernatant was removed, and then suspended in 50 mM Tris (pH 9.0), 3% BSA.

[0077] 2. Conjugation of Aminofluorescein Fluorescent Labeling Reagent

[0078] Colored polystyrene particles (see Table 1 and Figure 1~3 ) was diluted to 0.1%, and 6-aminofluorescein (Tokyo Chemical Industry Co., Ltd.) was added thereto so that the 6-am...

Embodiment 2

[0093] Example 2 Binding of Anti-Influenza A Virus NP Antibody and Fluorescent Labeling Agent to Visibly Colored Polystyrene Particles Having Carboxyl Groups as Functional Groups, and Detection of Influenza A Virus NP Antigen Based on Immunochromatography Using the Binding

[0094] In Example 2, polystyrene particles colored in the visible region were compared, and red polystyrene particles with high visibility and blue polystyrene particles I were used as materials for verification.

[0095] 1. Preparation of anti-influenza virus NP monoclonal antibody

[0096] BALB / c mice were immunized with type A influenza virus antigens, and spleens were removed from the mice that had been bred for a certain period of time. Mouse myeloma cells (P3×63) were fused. The resulting fused cells (hybridoma) were kept in a 37°C incubator, and the cells were purified (monoclonized) while confirming the antibody activity of the supernatant by ELISA using Influenza A virus NP antigen immobilized p...

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Abstract

The object of the present invention is to provide insoluble carrier particles used for high-sensitivity rapid immunoassay, which allow visual observation and high-sensitivity apparatus measurement. The insoluble carrier particles are visible colored insoluble carrier particles labeled with a fluorescent dye, which are used for immunoassay, wherein absorption of fluorescence by the visible colored insoluble carrier particles is low within a wavelength range of fluorescence emitted by the fluorescent dye.

Description

technical field [0001] The present invention relates to a method for specifically and rapidly detecting an analyte in a specimen (sample) and a kit using the same. Background technique [0002] Conventionally, immunoassays using insoluble carrier particles have been widely used. [0003] In the immunoassay method using insoluble carrier particles, the following method is adopted: on a test piece in which a ligand that specifically binds to a substance to be detected is immobilized at a predetermined position of a membrane having a fibrous structure such as a nitrocellulose membrane, using Gold colloid, platinum colloid, colored polystyrene particles, etc. are used as insoluble carrier particles, and the particles on which the ligand that specifically binds to the test substance is immobilized and the sample containing the test substance are developed on the above-mentioned surface by capillary phenomenon. On the test piece, the substance to be detected is interlayered with ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/543G01N21/64G01N33/545
CPCG01N21/6428G01N33/533G01N33/582G01N2021/6439
Owner DENKA CO LTD
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