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Ractopamine and cimaterolm combined colloidal gold test strip, and preparation method and application thereof

A technology of ractopamine and colloidal gold test paper, which is applied to the detection of β2-receptor agonist residues and the field of food safety, can solve the problems of large background interference, the need for improvement of ractopamine and cimaterol, and the inability to achieve quantitative detection. The effect of uniform size and fixed position

Inactive Publication Date: 2013-08-07
江西中德生物工程股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] (2) The sample matrix effect is obvious and the background interference is large
[0007] (3) Quantitative detection cannot be realized
[0008] (4) Only a single pollutant can be detected
[0010] Therefore, the current detection methods for the simultaneous quantitative detection of ractopamine and cimaterol still need to be improved.

Method used

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  • Ractopamine and cimaterolm combined colloidal gold test strip, and preparation method and application thereof
  • Ractopamine and cimaterolm combined colloidal gold test strip, and preparation method and application thereof
  • Ractopamine and cimaterolm combined colloidal gold test strip, and preparation method and application thereof

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Experimental program
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Embodiment 1

[0039] Synthesis of embodiment 1 immune antigen

[0040] 1.1 Synthesis of ractopamine immune antigen

[0041]Prepare ractopamine-BSA immunoantigen by mixed acid anhydride method: Weigh 34 mg of ractopamine and 10 mg of succinic anhydride to react in 2 mL of pyridine, stir overnight at room temperature, and place pyridine in a fume hood to completely evaporate. The reaction product is ractopamine-BSA Hemisuccinic anhydride; dissolve it in 2 mL of N,N-dimethylformamide and 2 mL of 1,4-dioxane mixture, add 26.2 μL of tri-n-butylamine, stir in an ice bath for 10 minutes, and then Add 15L of isobutyl chloroformate, react at room temperature, stir for 1 hour; add this mixture dropwise to a pre-cooled protein solution (100mg BSA dissolved in 0.1M sodium borate pH8.5), react overnight at room temperature, and dialyze in PBS for 72 hours Above, the purified ractopamine-BSA immune antigen can be obtained after dialysis.

[0042]

[0043] 1.2 Synthesis of cimaterol immune antigen

...

Embodiment 2

[0046] Example 2 Preparation and Titer Detection of Immunogen Monoclonal Antibody

[0047] 2.1 Preparation of ractopamine monoclonal antibody

[0048] Take 8-week-old female BALB / c mice, and use 0.1mL ractopamine-BSA and an equal volume of complete Freund's adjuvant emulsion to immunize each mouse for the first time by intraperitoneal injection; after that, take the same dose of immunogen to add With incomplete Freund's adjuvant, the same method was used to boost immunization once every month; the antibody titer was measured by indirect ELISA 10 to 14 days after the third and fourth immunizations, and finally the one with the highest antibody titer was selected for cell fusion. Spleen cells from immunized mice were taken under sterile conditions, fused with SP2 / O myeloma cells at a ratio of 5:1, added to HAT medium, 37 degrees Celsius, 6% CO 2 Cultivate in an incubator; replace half of the medium with fresh HAT medium after 5 days, and replace the HAT medium with HT medium af...

Embodiment 3

[0057] Embodiment 3 Preparation of ractopamine and cimaterol double colloidal gold test strips in the present invention

[0058] 3.1 Preparation of colloidal gold

[0059] The basic principle of preparing immune colloidal gold is that chloroauric acid can be polymerized into gold particles of a certain size under the action of a reducing agent to form a negatively charged hydrophobic colloid solution that is stable due to electrostatic interaction. The present invention adopts trisodium citrate reduction method to prepare colloidal gold, and specific process is as follows: take 0.01% chloroauric acid 100mL aqueous solution and heat to boiling, accurately add 1.5mL of 1% trisodium citrate aqueous solution under agitation, golden chloroauric acid is in Turn into purple within 2 minutes, turn off the heat source, continue to stir at high speed for 10 minutes, then reduce the speed to low gear, continue to stir for 1 hour, and restore the original volume with distilled water after...

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Abstract

The invention provides a ractopamine and cimaterolm combined colloidal gold test strip, and a preparation method and an application thereof. The test strip comprises a base plate with a first end and a second end. A filter paper, a sample pad, a gold label pad, a nitrocellulose membrane, and a water absorption pad are sequentially formed on the base plate from the first end to the second end. The gold label pad comprises colloidal-gold-labeled anti-ractopamine monoclonal antibody and colloidal-gold-labeled anti-cimaterolm monoclonal antibody. Two test lines and a control line are further formed on the nitrocellulose membrane. The test lines are sequentially composed of a ractopamine detection antigen linear point sample which can bind with the anti-ractopamine monoclonal antibody, and a cimaterolm detection antigen linear point sample which can bind with the anti-cimaterolm monoclonal antibody. The control line is composed of a donkey-anti-white-mouse antibody linear point sample which can bind with the anti-ractopamine monoclonal antibody and the anti-cimaterolm monoclonal antibody. With the cooperation of the test strip and a colloidal gold reading instrument, ractopamine and cimaterolm can be effectively and quantitatively detected.

Description

technical field [0001] The invention belongs to the field of food safety and involves β 2 - Field of detection of receptor agonist residues. Specifically, the present invention relates to a ractopamine-cimaterol dual-colloidal gold test strip and a preparation method and use thereof. Background technique [0002] Ractopamine and Cimaterol are both beta 2 - Receptor agonists. In recent years, many unscrupulous merchants have added them to animal feed as a substitute for clenbuterol hydrochloride. When used to feed animals for a long time, ractopamine and cimaterol are nutritionally beneficial to animals. The function of distribution promotes protein deposition in animals and inhibits fat formation, thereby significantly increasing the lean meat percentage of animal carcasses. However, ractopamine and cimaterol are metabolized slowly in the body, easy to accumulate, and have a high drug residue rate. When humans eat animal foods with residues of such drugs, they will be poi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/577G01N33/531
Inventor 赖卫华彭涛杨万春陈媛刘文娟
Owner 江西中德生物工程股份有限公司
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