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Anti-thiram monoclonal antibody, test paper for fast testing thiram and application thereof

A technology of monoclonal antibody and thiram, which is applied in the field of anti-thiram monoclonal antibody and detection of thiram composition, can solve the problems that affect the accuracy of analysis results, high technical requirements for instruments, expensive equipment and other problems, and achieve easy operation , low detection cost, and convenient storage

Inactive Publication Date: 2013-05-08
PEOPLES REPUBLIC OF CHINA BEIJING ENTRY EXIT INSPECTION & QUARANTINE BUREAU +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Chromatographic technology is very effective, accurate and sensitive for thiram detection, but it also has the following disadvantages: (1) sample pretreatment needs to be derivatized, and the procedure is cumbersome and time-consuming; (2) professional technicians are required to operate, and operators must be rich In order to obtain reliable analysis results, it is necessary to understand the various interference factors that affect chromatographic analysis and the advantages and disadvantages of the pretreatment method used; (3) It requires expensive instruments and equipment, and it is difficult for small and medium-sized cities and small and medium-sized enterprises to obtain reliable analysis results; (4) High technical requirements for instrument maintenance and maintenance
Whether the maintenance is proper will directly affect the accuracy of the analysis results; (5) The detection cost is high

Method used

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  • Anti-thiram monoclonal antibody, test paper for fast testing thiram and application thereof
  • Anti-thiram monoclonal antibody, test paper for fast testing thiram and application thereof
  • Anti-thiram monoclonal antibody, test paper for fast testing thiram and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0049] Example 1: Preparation of immunogold test paper for detecting thiram

[0050] 1. Preparation of thiram-carrier protein conjugate

[0051] 1) Weigh 40 mg of thiram and dissolve it in a 20mM PBS solution containing 0.02% Tween-20 to prepare a thiram solution.

[0052] 2) Under ice-water bath and magnetic stirring, add the Fumei double solution dropwise to 2.4ml 1mol / l HCl for diazotization (Itaru Y, Kohji I. Enzyme immunoassay forclenbuteol, an β 2 -adrnergicstimulant[J]. Journal of immunoassay, 1982, 3(2): 155-171) reaction, continue to react for 5 minutes after the addition.

[0053] 3) Under magnetic stirring, add the diazotized products dropwise to the human serum albumin (HSA), bovine serum albumin (BSA) and ovalbumin (OVA) solutions, and adjust the pH to 9.5 with 11mol / 1NaOH , Stir slowly overnight at 4°C to obtain three ligation products, namely thiram-HSA, thiram-BSA, and thiram-OVA.

[0054] 4) Put the ligation products thiram-HSA, thiram-BSA, and thiram-OVA obtained in s...

Embodiment 2

[0081] Example 2: Detection of sensitivity and specificity of anti-thiram monoclonal antibody

[0082] Coat the thiram-bovine serum albumin conjugate on the microwell plate, the coating amount is 10ng / well; use the anti-thiram monoclonal antibody and HRP enzyme to label to obtain the anti-thiram monoclonal antibody enzyme-labeled reagent , 1:1000 use; use PBST (0.5% Tween-20, 0.02MpH 7.4 phosphate buffer) as washing solution; use TMB color development system for color development; use 0.2M concentrated sulfuric acid for color development. Use this as a thiram detection system for testing, and the results are read by a microplate reader.

[0083] Choose buffer (0.01MpH7.4 phosphate buffer, formula: NaCl 8g, KCl0.2g, Na 2 HPO 4 ·12H 2 O 2.9g, KH 2 PO 4 0.2g, distilled water to make the volume to 1000ml), add thiram of different content and the concentration of 3000ppb thiram, ferram, thiram, propisen zinc, mancozeb, mancozeb, dithiocyanate Different types of samples are prepared wi...

Embodiment 3

[0088] Example 3: Detection of specificity and sensitivity of immune colloidal gold test paper

[0089] 1. Specific detection

[0090] The thiram, ferram, thiram, propsenzine, mancozeb, dithiocyanomethane, chlorothalonil, promethazine, and allicin were formulated into a sample with a concentration of 3000 ppb. Directly absorb 80 μl and add the immunogold test paper for detecting thiram of the present invention. The test results (see Table 2) show that the detection of thiram, ferram, thiram, propionate, mancozeb, dithiocyanomethane, chlorothalonil, promethazine, and allicin During the sample, a red band appeared on the test paper detection line and a red band appeared on the quality control line, indicating that thiram, thiram, thiram, propionate, mancozeb, dithiocyanate Methane, chlorothalonil, prodione, and ethicin have no cross-reaction with the test paper of the present invention, showing that the test paper of the present invention has good specificity.

[0091] Table 2: Resu...

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Abstract

The invention belongs to the field of pesticide detection and can fast detect the residual pesticide with an immunochemical technology. More particularly, the invention relates to a hybridoma cell line with the preservation number of CGMCC No.3580, and an anti-thiram monoclonal antibody generated by the hybridoma cell line. The invention further relates to a composite for detecting the thiram, a test paper for detecting the thiram, a preparation method of the test paper, an enzyme-coupled immunization agent for detecting the thiram, and application of the anti-thiram monoclonal antibody in detecting the thiram. The monoclonal antibody has high sensitivity and specificity. The test paper has the obvious advantages of high sensitivity, high specificity, convenient operation, fast and exact detection, etc.

Description

Technical field [0001] The invention belongs to the field of pesticide detection and uses immunochemical technology to quickly detect residual pesticides. Specifically, the invention relates to a hybridoma cell line and an anti-thiram monoclonal antibody produced by the hybridoma cell line. The present invention also relates to a composition for detecting thiram, a test paper for detecting thiram, a preparation method of the test paper, an enzyme-linked immunoassay reagent for detecting thiram, and the anti-thiram monoclonal antibody in the detection Application in Fumei Shuangzhong. Background technique [0002] Thiram, CAS registration number 137-26-8, chemical name is tetramethylthiuram disulfide, is a thiocarbamate pesticide, is a protective fungicide, its antibacterial spectrum Wide, can treat seeds and soil, prevent and cure cereal smut and seedling blight of various crops. After spraying, it can prevent some fruit trees and vegetable diseases. [0003] At present, the comm...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/577G01N33/558G01N33/544
Inventor 王金花罗海峰
Owner PEOPLES REPUBLIC OF CHINA BEIJING ENTRY EXIT INSPECTION & QUARANTINE BUREAU