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Malachite green collaurum detection card and production and application methods thereof

A malachite green, production method technology, applied in the field of immunology, to achieve the effect of low detection cost, cheap price and stable results

Active Publication Date: 2011-03-30
深圳市三方圆生物科技股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is no report on the monitoring of malachite green by this technology at home and abroad.

Method used

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  • Malachite green collaurum detection card and production and application methods thereof
  • Malachite green collaurum detection card and production and application methods thereof
  • Malachite green collaurum detection card and production and application methods thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] The synthesis of embodiment 1 malachite green hapten

[0043] Weigh 0.60 g of p-carboxybenzaldehyde, dissolve it in 20 mL of dichloromethane, and slowly add a 3-fold excess of thionyl chloride at room temperature. Gradually raise the temperature to reflux state, and react for 4h. The solvent and excess thionyl chloride were distilled off under reduced pressure. The crude product was dissolved in 20 mL of dichloromethane, cooled to 0°C, and 10 mL of 15% 4-aminobutyric acid aqueous solution was slowly added under rapid magnetic stirring. After 4 hours of reaction, the temperature was raised to room temperature and the reaction was continued for 2 hours. Adjust the pH to 6, and there is precipitation. The precipitate was filtered and washed twice with distilled water to obtain 0.35 g of the product 4-(4-formylbenzamido)butanoic acid.

[0044] Take 2.50g (20.0mmol) of N,N-dimethylaniline into a 50ml two-necked flask, slowly add 1.0-2.0ml of concentrated hydrochloric aci...

Embodiment 2

[0048] Synthesis of embodiment 2 malachite green immune antigen

[0049] Take malachite green hapten 0.1mmol and dissolve in 2mL DMF, stir and add 27.5mg DCC and 14.4mg NHS. Magnetic stirring was carried out overnight at 4°C. After centrifugation, the supernatant was liquid A, and 140 mg of BSA was weighed and dissolved in 10 mL of PBS (pH 8.0) with a concentration of 0.1 mol / L. Add 1 mL of DMF, stir and dissolve to prepare liquid B. Under magnetic stirring, liquid A is gradually dropped into liquid B, and react at 4°C for 12 hours. After centrifugation, the supernatant was taken, dialyzed with normal saline at 4°C for 3 days, and the dialysate was changed 3 times a day. The obtained whole antigen was dispensed into 0.5 mL centrifuge tubes at a concentration of 1 mg / mL. Store in -20°C freezer.

[0050] The prepared immune antigen structure is:

[0051]

Embodiment 3

[0052] Embodiment 3 experimental animal immunization

[0053]Female balb / c mice were immunized with the hapten malachite green immunization antigen for 6 weeks, 3 mice in each group. For the first immunization injection, 100 μL of 100 μg / mL immune antigen was fully emulsified with the same amount of complete Freund’s adjuvant, and injected directly into the intraperitoneal cavity. After an interval of two weeks, take the sampled antigen, emulsify it with 100 μL of incomplete adjuvant, and inject it in the same way.

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Abstract

The invention relates to a malachite green collaurum detection card and production and application methods thereof, belonging to the technical field of immunology. The rapid diagnosis card comprises a case and a test strip; wherein the case is opened with a sample injecting hole and an observation window, the test strip is arranged in the case, the middle part of a PVC liner plate is superposed with a nitrocellulose membrane, the two ends are respectively superposed with an absorbent pad and a sample pad, the nitrocellulose membrane is overlapped and connected with the absorbent pad and the sample pad, a membrane containing malachite green monoclonal antibody immune collaurum compound is coated on a sample region, and a detection strip containing malachite green envelop antigen and a quality control strip containing goat anti mouse IgG are arranged on the nitrocellulose membrane. The invention is a rapid, accurate and sensitive diagnosis card, requires no special apparatus, has simple operation and can meet the detection requirements of food safety, fishery and detection mechanism.

Description

technical field [0001] The invention relates to a malachite green colloidal gold detection card, which belongs to the technical field of immunology. Background technique [0002] Malachite green is widely used as a fungicide in fishery, which brings great harm. Malachite green is a kind of green crystal with metallic luster, also known as alkaline green, strict block green, malachite green, it is not only a fungicide, but also a dye, easily soluble in water, and the solution is blue-green; soluble in Methanol, ethanol and pentanol. For a long time, fishermen have used it to prevent fish saprolegniasis, gill mold disease, small melon worm disease, etc., and in order to prolong the life of fish with damaged scales, malachite green is also often used during transportation and in storage pools. . Scientific research results show that malachite green remains in fish for a very long time, and it has high toxin, high residue and side effects such as carcinogenic, teratogenic and...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/577G01N33/558G01N33/543C07K14/765C07K14/77
Inventor 钟松清谭攀李细清张世伟
Owner 深圳市三方圆生物科技股份有限公司
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