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Immuno colloidal gold test paper strip for detecting sulfa drug residue

The technology of sulfonamide drugs and test strips is applied in the field of immunochemical rapid detection of veterinary drug residues, which can solve the problems of complex operation process, high requirements for instrument maintenance, and high detection costs, and achieves multi-functionality and good specificity, and storage temperature. The effect of low requirements and short detection time

Inactive Publication Date: 2008-06-25
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] 1. Chromatographic technology is very effective, accurate and sensitive for the detection of sulfa drugs, but it also has the following disadvantages: (1) The pretreatment procedure of the sample to be tested is cumbersome and time-consuming (2) It needs professionally trained technicians to operate, and the operators need to With rich relevant experience, the operator must understand the various interference factors affecting chromatographic analysis and the advantages and disadvantages of the pretreatment method used in order to obtain reliable analysis results; limit
(4) The requirements for instrument maintenance are high, and the quality of maintenance directly affects the accuracy of the analysis results; (5) The detection cost is high
The disadvantages are: (1) special instruments and equipment such as microplate reader are required to be used together, which is not conducive to popularization and use in grassroots units; (2) detection operators need to undergo professional training; (3) the operation process is relatively complicated, The time required for the detection is relatively long, and the whole process takes 2h-4h; (4) The cost required for the detection is relatively high, and a single detection cannot be realized

Method used

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  • Immuno colloidal gold test paper strip for detecting sulfa drug residue
  • Immuno colloidal gold test paper strip for detecting sulfa drug residue
  • Immuno colloidal gold test paper strip for detecting sulfa drug residue

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040] Embodiment 1 (preparation embodiment)

[0041] Preparation method of immunocolloidal gold test strip for detecting sulfa drugs

[0042] 1. Synthesis of the small molecular hapten N-sulfa-4-aminobenzoic acid (SDL), which contains p-aminobenzenesulfonamide, the common core structure of sulfa drugs. The specific steps are:

[0043] (1) Accurately weigh 3.2 g of methyl p-aminobenzoate, dissolve it in 20 mL of anhydrous pyridine, and pour it into a 100 mL two-neck round bottom flask.

[0044] (2) While stirring, slowly add 6.0 g of vacuum-dried p-acetamidobenzenesulfonyl chloride into the round bottom flask, and after the addition, reflux on the oil bath to make the solution turn red.

[0045] (3) After refluxing for a few minutes, the reaction solution turned into a very viscous red substance, and after a few minutes the reaction solution turned into a brown liquid, and continued to reflux for 15 minutes, and the reaction solution turned into a brownish-yellow liquid. Af...

Embodiment 2

[0079] Embodiment 2 (application embodiment)

[0080] The use method of the immunocolloidal gold test strip for detecting sulfa drugs of the present invention

[0081] 1. Sample pretreatment

[0082] (1) Pretreatment of tissue samples

[0083] Homogenize the tissue sample to be tested, take 2 g of the tissue sample to be tested, add 4 mL of ethyl acetate, shake up and down for 5 min, centrifuge at 2000 g at room temperature for 5 min, take 300 μL of the upper layer liquid and evaporate it to dryness in a water bath at 80 ° C or dry it under nitrogen flow, and use 150 μL of pH7.4 Phosphate buffer (recipe: NaCl 8g, KCl 0.2g, NaCl 2 HPO 4 12H 2 O 2.9g, KH 2 PO 4 0.2 g, dilute to 1000 mL with distilled water) to dissolve the residue.

[0084] (2) Pretreatment of milk samples

[0085] After the milk sample was degreased by centrifugation at 3500rpm for 10min, absorb 2mL of the middle layer of milk and add 4mL of ethyl acetate to shake up and down for 5min, centrifuge at 200...

Embodiment 3

[0092] Embodiment 3 (application embodiment)

[0093] Application of the immunocolloidal gold test strip for detecting sulfa drugs of the present invention

[0094] 1. Specificity test

[0095] According to the method described in Example 2, it is 10, 20, 40, 80, 100 ng / mL sulfamethoxine and the negative standard substance that are respectively detected with the detection test strip of the present invention, and are measured with the Biodot TSR3000 strip reading system. The detection line absorbance value (G / Peak-ROD value), taking the concentration of sulfamethoxine as the X axis, and the detection line absorbance value (G / Peak-ROD value) as the Y axis draw a standard curve, and calculate the sulfamethoxine according to the standard curve The 50% inhibitory concentration of oxypyrimidine was 18.3ng / mL. Similarly, the 50% inhibitory concentration of sulfamethoxine, sulfadimethoxine, sulfadiazine, sulfaquinoxaline, sulfamethazine, sulfamethoxazole, and trimethoprim was calcul...

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Abstract

The present invention belongs to the field of veterinary medicine residue detecting immunochemistry technology, and discloses one kind of colloidal gold test paper strip for fast detection of sulfa drug residue. The test paper strip consists of sample pad, combining pad, nitrocellulose film, water absorbing pad and PVC back lining. It features the combining pad with coated sulfa drug resisting specific monoclonal antibody-colloidal gold marker, the nitrocellulose film with coated N-sulfanilamino-4-amino benzoic acid-carrier protein coupler as the detection line and coated rabbit antimouse IgG as quality control line. The present invention also discloses the preparation of hybridoma BDRXN-2 with preservation number of CCTCC-C200629 capable of secreting specific sulfa drug recognizing monoclonal antibody. The test paper strip of the present invention has the advantages of high sensitivity, high specificity, simple operation, fast detection and low cost.

Description

technical field [0001] The invention belongs to the technical field of immunochemical rapid detection of veterinary drug residues, and specifically relates to an immunochromatographic reaction using colloidal gold markers for color development, and an immunocolloidal gold test strip for rapidly detecting sulfonamide drug residues. Background technique [0002] Since my country's accession to the WTO, people have paid more and more attention to food safety issues. The government has never paid more attention to food safety issues. Among them, the residual pollution of agricultural and veterinary drugs in food and agricultural by-products is the most concerned safety and health issue at home and abroad. [0003] Sulfonamides are artificially synthesized broad-spectrum antibacterial drugs with the core structure of p-aminobenzenesulfonamide. As veterinary drugs and feed additives, they are widely used in the feeding of food-borne animals and have significant effects in the preve...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/18C07K16/00G01N33/577G01N33/52
Inventor 毕丁仁王喜亮李奎熊宁石德时刘百红彭伏虎郝卫芳金秀娥
Owner HUAZHONG AGRI UNIV
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