Rapid immunomagnetic bead chromatographic method for main allergens in aquatic products

A main allergen and rapid immunization technology, applied in the field of food allergen detection methods, to achieve strong specificity, easy operation, and high sensitivity

Active Publication Date: 2011-05-04
SHANGHAI OCEAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0013] At present, there are at least 14 commercial immunochromatographic test kits on the market for detecting allergens including milk, peanuts, hazelnuts, and crustacean aquatic products, but these products use colloidal gold immunochromatography, which can only be qualitative Or semi-quantitative monitoring. At present, there is no rapid on-site detection technology that can quantitatively detect the main allergens of aquatic products at home and abroad.

Method used

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  • Rapid immunomagnetic bead chromatographic method for main allergens in aquatic products
  • Rapid immunomagnetic bead chromatographic method for main allergens in aquatic products
  • Rapid immunomagnetic bead chromatographic method for main allergens in aquatic products

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preparation example Construction

[0051] The preparation method of the specific nanometer detection probe is:

[0052] A. Take an appropriate amount of carboxyl magnetic beads into a centrifuge tube, use 500μL 0.01M MEST solution containing 0.5% (v / v) Tween-20, pH5.0 as the activation buffer solution to wash the magnetic beads, and place the centrifuge tube on the magnetic separation rack Make the magnetic beads separate from the activation solution, repeat the washing several times, and finally resuspend the magnetic beads;

[0053] B. Then add freshly prepared 2.6M carbodiimide (EDC) and 2.2M N-hydroxysuccinyl

[0054] Add imine (NHS) to the magnetic bead suspension to activate the carboxyl group for 30 minutes, then wash the magnetic beads with MEST buffer; then wash the magnetic beads twice with 0.005M borate Tween (abbreviated as BST) solution as the coupling buffer;

[0055] C, add parvalbumin monoclonal antibody subsequently, react 3h on the rotary mixer;

[0056] D. Then use 1% w / v BSA solution (BSA ...

Embodiment 1

[0065] Embodiment 1: Preparation of the main allergen Tm rapid detection test strip of aquatic products

[0066] In this embodiment, Tm is used as the detection object, and magnetic nanoparticles (particle size: 200nm) modified with carboxyl groups on the surface are selected. The Tm that can detect crustacean edible aquatic organisms such as shrimp, crab and clam can also be used to detect conch, abalone, The mouse monoclonal specific reaction to Tm allergens in edible aquatic organisms such as squid and octopus was used to prepare specific probes for Tm allergens in seafood, and the detection line was designed using the competitive reaction principle.

[0067] (1) Preparation of specific nano-detection probes for allergen Tm

[0068] EDC / NHS coupling

[0069] The Tm-specific monoclonal antibody was coupled with carboxyl-modified nano magnetic beads (particle size: 200nm) to prepare immunomagnetic beads containing Tm monoclonal antibody, that is, Tm magnetically label...

Embodiment 2

[0079] Embodiment 2: Preparation of the main allergen Pa rapid detection test strip of aquatic products

[0080] In this example, Pa is used as the detection object, and magnetic nanoparticles (particle size: 100nm) modified with carboxyl groups on the surface are selected, combined with those that have specific reactions to parvalbumin in fish and Pa in vertebrates such as frogs, rats, and turtles. Murine monoclonal antibodies were used to prepare specific nanoprobes for the main fish allergen Pa, and then the detection line was designed using the competitive reaction principle.

[0081] (1) Preparation of specific nanoprobes for allergen Pa

[0082] EDC / NHS coupling

[0083] Coupling the parvalbumin monoclonal antibody with carboxyl-modified nano-magnetic beads to prepare immunomagnetic beads containing parvalbumin monoclonal antibody, that is, parvalbumin magnetic-labeled antibody. Use MEST solution (0.05% Tween-20) at pH 5.0 as the activation buffer solution, take...

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Abstract

The invention relates to a rapid immunomagnetic bead chromatographic method for main allergens in aquatic products. A rat monoclonal antibody which has specific reaction with a tropomyosin (Tm) allergen of edible crustacean aquatic organisms and edible soft-bodied aquatic organisms, has specific reaction with a parvalbumin (Pa) allergen of fishes, frogs, rats and sea turtle vertebrates, and is prepared and screened by hybridoma technology is taken as a specific antibody and coupled to the surface of a magnetic nanometer material in a chemical bonding mode to serve as a marker so as to prepare a specific nanometer detection probe suitable for magnetic bead chromatography; the allergen specific nanometer detection probe and sample solution are added into a sample pad of an allergen magnetic immunochromatographic test strip, the sample pad is stood at room temperature for 20 minutes, the allergens are captured by a detection strip and a control strip through chromatography to form a macroscopic colored strip, so that immunochromatography is realized; and a test card is put on a magnetic assay reader, the content of the allergens in a sample to be detected is calculated according to a marked line, comparison is performed and the immunochromatography is finished.

Description

technical field [0001] The present invention relates to the detection method of food allergen, relate in particular to an immunomagnetic bead chromatography method for detecting main allergens in aquatic products, the method mainly uses the main allergen myosin (Tropomyosin) , referred to as Tm), the main fish allergen parvalbumin (Parvalbumin, referred to as Pa) immunomagnetic bead chromatography detection method. Background technique [0002] Food allergy is an abnormal response of the body to the immune system caused by food. In recent decades, the number of patients with food anaphylaxis has been increasing year by year worldwide. According to WHO reports, by 2010, there may be 40% to 50% of the global population. People suffer from allergies, and food allergy is now an emerging food safety issue. Food allergens are substances in food that trigger or provoke an allergic reaction, usually naturally occurring proteins that are abundant in certain foods. According to the ...

Claims

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Application Information

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IPC IPC(8): G01N33/577G01N33/531
Inventor 卢瑛王锡昌郑诚石良
Owner SHANGHAI OCEAN UNIV
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