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Method for detecting by using quantum dot fluorescence immunochromatographic test strips

A fluorescence immunochromatography and quantum dot technology, applied in the field of immunodetection, can solve the problems affecting detection sensitivity and poor precision, and achieve the effects of good labeling stability, simple steps and safe operation.

Active Publication Date: 2013-04-17
WUHAN JIAYUAN BIOMEDICAL ENG
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  • Abstract
  • Description
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AI Technical Summary

Problems solved by technology

However, various existing immunochromatographic test strip detection methods involving quantum dots are either due to the selection of nuclear quantum dots, quantum dots synthesized in water or quantum dot-coated microspheres, etc., or due to the adoption of a solidification method to It is fixed in the binding area, which affects the improvement of detection sensitivity and causes poor precision

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  • Method for detecting by using quantum dot fluorescence immunochromatographic test strips
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Examples

Experimental program
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Effect test

Embodiment 1

[0023] Example 1: Diagnosis of high-sensitivity C-reactive protein

[0024] Preparation of quantum dot-labeled protein detection solution: the carboxyl water-soluble quantum dots used are carboxyl water-soluble core-shell quantum dots. The emission wavelength range of quantum dots is 500nm-630nm, which can emit fluorescence of different wavelengths under the action of excitation light source radiation. Its fluorescence quantum yield is not less than 45%. Take 250 μL (2nmol) of carboxyl water-soluble quantum dots with a concentration of 8 μM, equilibrate to room temperature, and treat with ultrasonic wave 200W for 10 seconds, add 57.5 μL of 10 mg / mL EDC (carbodiimide hydrochloride) according to the ratio of 3 μmol EDC / 2nmol quantum dots , vortexed to mix, and then added 10 mg / mL antibody 0.6mL according to the ratio of 40nmol anti-human C-reactive protein monoclonal antibody / 2nmol quantum dots, and vortexed to mix. Quantum dots, EDC, protein, etc. were all dissolved with 20mM~...

Embodiment 2

[0030] Embodiment 2: Detection of aflatoxin M1

[0031] Preparation of quantum dot-labeled protein detection solution: Take 250 μL (2 nmol) of carboxyl water-soluble quantum dots with a concentration of 8 μM, equilibrate to room temperature, and treat with ultrasonic wave 200W for 10 seconds, add 10 mg / mL EDC (carbon Diimine hydrochloride) 57.5 μL, vortexed to mix, then add 10mg / mL antibody 0.6mL according to the ratio of 40nmol anti-aflatoxin M1 monoclonal antibody / 2nmol quantum dots, vortexed to mix. Quantum dots, EDC, protein, etc. were all dissolved with 20mM~200mM pH6.0~7.4 phosphate solution, and 1.1mL phosphate solution was added to the above solution system. The reaction was stirred at room temperature for 1-4 hours. After the reaction, use a centrifugal ultrafiltration tube with a molecular weight cut-off of 50 kDa to remove unreacted impurities and concentrate to 20 μL, then purify through a gel separation column filled with Superdex G200 filler, collect the fluores...

Embodiment 3

[0037] Example 3: Realization of joint detection of HFMD-associated Coxsackievirus A16 and Enterovirus 71

[0038] Preparation of quantum dot-labeled protein detection solution: Refer to Example 1, using green and orange-red quantum dots with emission wavelengths of 525nm and 605nm to label Coxsackievirus A16 (CA16) monoclonal antibody and enterovirus 71 (EV71) respectively Monoclonal antibody, mouse IgG is labeled with yellow quantum dots with an emission wavelength of 565nm, purified and concentrated to 1-2μM, mixed with the same concentration of the three labeled proteins, diluted 100-1000 times with PBS-TBN after purification, in 2-8 ℃ refrigerator for future use.

[0039] Assembly of immunochromatographic test strips: as figure 2 As shown, the reaction film was fixed and pasted on the middle part of the PVC backing, and the concentration of the antibody and goat anti-mouse IgG antibody were respectively adjusted to 1 mg / mL and 0.2 mg / mL with coating buffer, and Coxsacki...

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Abstract

The invention provides a method for detecting by using quantum dot fluorescence immunochromatographic test strips. The method comprises the following specific steps: (1) preparing a quantum dot labeled protein liquid; (2) preparing a quantum dot labeled protein detecting liquid; (3) preparing a reaction film coated with a detection index; (4) bonding a sample pad, the reaction film prepared in the step (3) and a water absorbing pad on a PVC backing in sequence so as to obtain a test paper board; cutting the test paper board into test strips; and loading the test strips into a test paper clip; and (5) qualitatively or quantitatively detecting. The method for detecting by using the quantum dot fluorescence immunochromatographic test strips, provided by the invention, is a rapid detecting method using the quantum dot fluorescence immunochromatographic test strips, which is featured by high sensitivity, synchronicity in multi-index detection, simplicity, convenience, intuitive nature, low price and wide application.

Description

technical field [0001] The invention relates to a detection method using quantum dot fluorescent immunochromatographic test strips, which belongs to the field of immune detection. Background technique [0002] Immunochromatography test strip detection, also known as immunological lateral flow chromatography, is a fast, simple, sensitive, intuitive, inexpensive, and on-site detection method. It has the advantages that radioimmunoassay, enzyme-linked immunosorbent assay, gas chromatography, liquid chromatography, capillary electrophoresis and other instrument detection methods do not have. It plays an important role in detection technology and is a powerful supplement to traditional and large-scale instrument detection methods. . With the development of society, human beings have higher requirements for the quality of life. Immunochromatographic detection technology plays an important role in the face of major issues such as human diseases, environmental pollutants, food safe...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/577
Inventor 彭俊朱小波庞代文张志凌余慧
Owner WUHAN JIAYUAN BIOMEDICAL ENG
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