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Kit and method for highly sensitive detection of allergen-specific antibody ige

A sensitive detection and allergen technology, applied in the direction of biological testing, material inspection products, etc., to achieve the effect of less sample consumption

Active Publication Date: 2016-09-28
ZHEDA DIXUN BIO GENE ENG
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

For example, ImmunoCAP Rapid, a product of Pharmacia, Sweden, can only screen a dozen kinds of allergens at a time, without a reader, and the sensitivity of naked eyes to judge the concentration of sIgE antibody is only 1.0IU / ml (1IU IgE=2.44ngIgE), and 1.49IU / ml is used to distinguish negative and a positive result

Method used

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  • Kit and method for highly sensitive detection of allergen-specific antibody ige
  • Kit and method for highly sensitive detection of allergen-specific antibody ige
  • Kit and method for highly sensitive detection of allergen-specific antibody ige

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040] Embodiment 1: detection reagent preparation of the present invention

[0041] 1. Nitrocellulose membrane immobilized (covalently bound) allergen protein (refer to Hermanson GT et al, Immunobilized affinity ligand techniques. Academic Press (New York), 1992, pp53-56.)

[0042] (1) nitrocellulose membrane activation

[0043] A. After wearing the safety protection equipment, prepare acetonitrile solution of CNBr (US Sigmaaldrich Company, C91492) with a concentration of 1mg / ml in the fume hood;

[0044] B. Suspend the cut nitrocellulose membrane (Bio-Rad, USA, 162-0112) in 1mol / L, pH11 Na 2 CO 3 solution, transferred to a glass beaker with a stirring bar, and then placed flat on a magnetic stirrer plate in a fume hood;

[0045] C. While stirring, add CNBr solution; stir for 10 minutes, during which the pH value of the solution is measured with precision pH test paper above pH 8 every 2 minutes; if necessary, adjust the pH of the activation solution to 11 with NaOH soluti...

Embodiment 2

[0068] Example 2 (signal amplification system without using biochemical methods):

[0069] For the preparation of detection reagents, refer to Example 1 (excluding dual-free biotin-streptavidin optimization solution and biotin-labeled poly-alkaline phosphatase solution).

[0070] The detection method is as follows:

[0071] (1) Pretreatment: Take out the detection reaction tank required for the experiment (for the structure, see image 3 , the fibrous material film solidified with the allergen protein to be tested has been installed in the small hole), placed horizontally at room temperature, numbered or marked with the patient's name;

[0072] (2) Initial incubation: Add 300 microliters of serum to 1.7ml of sample diluent (PBS at pH 7.4 + 10% newborn bovine serum) and mix well, pour it into the reaction tank with the cover film removed, and place the reaction tank in On a mixer, incubate at room temperature (20-25°C) for 45 minutes.

[0073](3) Cleaning: Rinse the liquid r...

Embodiment 3

[0081] Example 3 (secondary signal amplification system using dual biotin-streptavidin and polymerase):

[0082] For the preparation of detection reagents, see Example 1.

[0083] The detection method is as follows:

[0084] (1) Pretreatment: Take out the detection reaction tank required for the experiment (for the structure, see image 3 , the fibrous material film solidified with the allergen protein to be tested has been installed in the small hole), placed horizontally at room temperature, numbered or marked with the patient's name;

[0085] (2) Initial incubation: Add 300 microliters of serum to 1.7ml of sample diluent (PBS at pH 7.4 + 10% newborn bovine serum) and mix well, pour it into the reaction tank with the cover film removed, and place the reaction tank in On a mixer, incubate at room temperature (20-25°C) for 45 minutes.

[0086] (3) Cleaning: Rinse the liquid reaction tank with washing liquid (PBS at pH 7.4 + 0.05% Tween-20), and repeat washing 5 times for 10...

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Abstract

The invention provides a kit and a method for detecting specific allergen antibody IgE in a high-sensitivity manner. The kit mainly comprises a fiber material membrane provided with allergen protein to be detected, double biotin-streptavidin optimized liquid, a biotin coupled anti-human IgE antibody, biotin or streptavidin marked polymerase, and a substrate color developing agent corresponding to polymerase in a curing manner; the kit can qualitatively or semi-quantitatively detect the concentration of the specific allergen antibody IgE in human serum or plasma rapidly in the high-sensitivity manner, can screen dozens of allergens, is rapid and accurate, and is suitable for high-throughput testing, and a relatively small quantity of samples are used.

Description

(1) Technical field [0001] The invention relates to a highly sensitive detection kit for qualitatively or semiquantitatively screening or detecting multiple allergen-specific IgE antibody levels in human serum or plasma, and a detection method thereof. (2) Background technology [0002] From 2011 to 2012, the World Allergy Organization (WAO) white paper on allergy (also known as anaphylaxis, Allergy) pointed out: "The prevalence of allergic diseases is rising dramatically worldwide in developed and developing countries. Allergic Diseases include allergic asthma, allergic rhinitis, severe allergic reactions, allergic reactions to drugs, food and insects, eczema, urticaria (hay fever) and angioedema. The increase in the number of allergic diseases, especially among children, in the past two decades has Outbreak trends.” (Pawankar R et al, WAO White Book on Allergy 2011-2012: Executive Summary.) The prevalence of allergic diseases caused by food alone is 3% to 6%. (Sicherer S ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/68
CPCG01N33/68
Inventor 臧荣春吴善东
Owner ZHEDA DIXUN BIO GENE ENG
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