A method for detecting the specific immunoglobulin e of Penaeus lanceolata

An immunoglobulin and detection method technology, which is applied in the field of detection of specific immunoglobulin of Penaeus knives, can solve the problems of high detection cost, interference of allergy detection results, and inability to standardize the sensitization efficiency.

Active Publication Date: 2019-04-23
JIANGNAN UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

But it is also one of the eight types of highly allergenic foods, with an incidence rate of 15.6%-35%, leading to the occurrence of allergic asthma, anaphylactic shock and other diseases
[0003] At present, my country is in its infancy in the field of allergy-related research, and there are no relevant diagnostic reagents for shrimp and crab food allergies
Clinical detection reagents mainly rely on imports, such as the Immuno CAP system automatic detection system certified by the US FDA, its recognized detection limit is 0.24ng / mL, and the detection cost is relatively expensive, most allergic patients cannot afford it; The wide range of ELISA detection kits has a high detection limit and cannot accurately identify whether they are allergic patients
Moreover, the allergens used in allergy testing are crude extraction products of biological materials. The crude extraction products contain allergens, non-specific allergens, and suspected allergens that have not yet been characterized, which interfere with the results of allergy testing, and their sensitization efficiency cannot be standardized.

Method used

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  • A method for detecting the specific immunoglobulin e of Penaeus lanceolata
  • A method for detecting the specific immunoglobulin e of Penaeus lanceolata
  • A method for detecting the specific immunoglobulin e of Penaeus lanceolata

Examples

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Embodiment 1

[0031] Example 1: Preparation of fluorescent probes with different particle sizes FHMNs@PDDA@PAA

[0032] Dissolve cetyltrimethylammonium bromide (CTAB) powder in a mixture of 29mL deionized water, 12mL ethanol and 1mL ammonia water, then add polystyrene microsphere dispersion (dispersed in 10ml deionized ionized water), after magnetically stirring at room temperature for 60 min, tetraethyl orthosilicate (TEOS) was added dropwise, maintaining a constant stirring speed of 150 rpm / min, and continuing to stir at room temperature for 48 h. After the reaction was completed, the reaction solution was vacuum filtered and washed several times, and then dried in a vacuum oven at 65°C for 12 hours to obtain core-shell PS / SiO 2 Composite microsphere powder. Calcined at 550°C for 8h to remove template polystyrene to obtain hollow silica. Disperse 0.05g of HMNs and 0.012g of 5(6)-fluorescein isothiocyanate (FITC) in 5ml of deionized water, stir at room temperature for 48h, centrifuge the...

Embodiment 2

[0034] Example 2: Magnetic probe Fe 3 o 4 @SiO 2 Preparation of @PAA Nanoparticles

[0035] Weigh 300mgFe 3 o 4 Disperse in a mixture of 40mL ethanol and 4mL deionized water, after ultrasonication for 15min, maintain a certain stirring speed, add 5mL ammonia water, slowly add 2mLTEOS, react at room temperature for 12h, and use 0.1moL / L hydrochloric acid solution and deionized water after magnetic separation Wash each for 3 times, and dry at 40° C. for 12 hours to obtain a reddish-brown precipitate, which is ferric oxide / silicon dioxide core-shell nanoparticles. Fe will be produced 3 o 4 @SiO 2 Dispersed in dimethylamide (DMF), mixed with 33ml DMF dissolved in 2g polyacrylic acid (PAA), ultrasonicated for 30min, the mixture was vigorously stirred and heated to 110°C, added 3.3mL DMF dissolved in 4-dimethylaminopyridine and Dissolve N,N'-dicyclohexylcarbodiimide (DCC) in 6.6mL DMF, keep the mixture at 110°C and stir for 12h, then magnetically separate the product, wash w...

Embodiment 3

[0037] Example 3 pH optimization process, time optimization process, and temperature optimization process respectively provide pH optimization

[0038] Take appropriate amount of Fe 3 o 4 @SiO 2 @PAA—antigens and FHMNs@PDDA@PAA—Ab2 were added with positive IgE (specific immunoglobulin E) of Penaeus lanceolata, and after incubation at a certain temperature, the precipitate was collected by magnetic separation and washed with phosphate buffer (pH7.4) Wash 3 times, collect the precipitate by magnetic separation, add 3mL sodium hydroxide solution of different pH and sonicate for 20min, dissolve the hollow mesoporous shell, release fluorescent molecules, and detect the fluorescence intensity, the results are as follows: Figure 4 As shown in A, the fluorescence intensity is the highest at pH 11, so the sodium hydroxide solution at pH 11 is selected as the detection solution.

[0039] temperature optimization

[0040] Take appropriate amount of Fe 3 o 4 @SiO 2 @PAA—antigens a...

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Abstract

The invention discloses a method for detecting specific IgE (immunoglobulin E) of metapenaeus ensis and belongs to the cross field of materials and biomedicine. FHMNs (fluorescent hollow mesoporous silica nanoparticles) are adopted as a vector of an anti-IgE antibody, Fe3O4@SiO2 prepared through coating of Fe3O4 with SiO2 is used as an allergen vector, the two materials capture targeted sIgE (specific IgE) of the metapenaeus ensis simultaneously, the targeted sIgE is quickly enriched and separated by use of magnesium of Fe3O4@SiO2, the signal of low-concentration targeted sIgE is amplified by use of fluorescent molecular signals in the FHMNs, the fluorescence intensity is associated with concentration of the IgE, and the sIgE of the metapenaeus ensis in human blood is detected quickly and sensitively. With the adoption of the method, the IgE detection limit is effectively reduced and the detection time is shortened.

Description

technical field [0001] The invention relates to a method for detecting the specific immunoglobulin E of Penaeus lanceolata, which belongs to the intersection field of materials and biomedicine. Background technique [0002] Shrimp and crab are the most common crustaceans in my country's aquatic product market, and they are high-protein and high-nutrition foods. But it is also one of the eight types of highly allergenic foods, with an incidence rate of 15.6%-35%, leading to the occurrence of allergic asthma, anaphylactic shock and other diseases. [0003] At present, my country is in its infancy in the field of allergy-related research, and there are no relevant diagnostic reagents for shrimp and crab food allergy. Clinical detection reagents mainly rely on imports, such as the Immuno CAP system automatic detection system certified by the US FDA, its recognized detection limit is 0.24ng / mL, and the detection cost is relatively expensive, most allergic patients cannot afford ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/68G01N33/533C09K11/02C09K11/06
CPCC09K11/025C09K11/06G01N33/533G01N33/6854G01N2333/47
Inventor 吴静云建荣姚瑞
Owner JIANGNAN UNIV
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