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Kit for rapidly detecting content of dalapon in crops

It is a technology of cod and kit, which is applied in the field of monoclonal antibody preparation and ELISA kit, specific monoclonal antibody, and ELISA. Measure a large number of samples and other problems to save time and reduce operation steps

Inactive Publication Date: 2016-01-27
JIANGSU WISE SCI & TECH DEV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, HPLC is the main method for the determination of thatraquat. Due to the shortcomings of this method, such as long analysis time, large sample volume required for detection, and cumbersome pretreatment steps, it is not suitable for the determination of a large number of samples.

Method used

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  • Kit for rapidly detecting content of dalapon in crops

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] The preparation of embodiment 1 kit components

[0044] 1. Antigen synthesis

[0045] a. Dissolve 10g of carrier bovine serum albumin (BSA) in 35mL of pH9.6 carbonate buffer, add 10g of ethylenediamine to activate;

[0046] b. Dissolve 1 g of palapaquat in 10 mL of 0.5 M sodium hydroxide solution;

[0047] c. Take 1g of carbodiimide and dissolve it in 5mL of pure water, then add it to the solution of palapaquat and stir for 3 hours at room temperature;

[0048] d. Add the activated carrier protein BSA dropwise to the palapaquat solution and stir at 4°C overnight;

[0049] e. The artificial antigen obtained after the reaction was dialyzed against 0.1M PBS buffer for 5 days, and the buffer was changed 4 times a day; the obtained purified artificial antigen was concentrated by ultrafiltration or lyophilized for storage.

[0050] 2. Preparation of monoclonal antibody against thatraquat

[0051] a. Animal immunization procedure: use the artificial antigen of palapaquat...

Embodiment 2

[0066] Embodiment 2 is used for the formation of the enzyme-linked immunoimmunoassay kit of rapid detection of palapa

[0067] This ELISA kit consists of the following components:

[0068] (1) A microtiter plate coated with the palapaquat antigen;

[0069] (2) Malawat monoclonal antibody working solution with a protein concentration of 0.5 μg / L;

[0070] (3) A standard substance with a concentration of 1 μg / mL of thatraquat;

[0071] (4) Standard diluent: 0.05mol / LTris-HCl, pH8.0, 0.9%NaCl buffer;

[0072] (5) Goat anti-mouse enzyme-labeled secondary antibody working solution labeled with horseradish peroxidase;

[0073] (6) Hydrogen peroxide at the liquid level of substrate chromogenic solution A, and o-phenylenediamine at the liquid level of substrate chromogenic solution B;

[0074] (7) The washing buffer is 0.05mol / LTris-HCl, pH8.0, 0.9%NaCl, 0.04%Tween20;

[0075] (8) The stop solution is 2mol / L sulfuric acid.

Embodiment 3

[0076] The detection of thatraquat residue in the sample of embodiment 3

[0077] 1. Sample pretreatment

[0078] a. Take 1g of pulverized and representative sample, add 5mL of acetone;

[0079] b. Fully oscillate and mix for 3-5 minutes, after standing for stratification, take 5mL supernatant and centrifuge at 4000r / min for 5min or filter with quantitative analysis filter paper;

[0080] c. Take 1 mL of the centrifuged supernatant or filtered filtrate, and dry it with nitrogen in a water bath at 50-60 °C;

[0081] d. Add 1mL of standard diluent and mix well, centrifuge at 4000r / min for 5min or filter with quantitative analysis filter paper;

[0082] e. Take the centrifuged supernatant or filtered filtrate for analysis.

[0083] 2. Detection with kit

[0084] Prepare a mother solution with a concentration of 1000ng / mL palapaquat, perform double dilution, and dilute to 0, 0.3, 0.9, 2.7, 8.1, 24.3ng / mL 6 concentrations of standard products, according to 100μL standard or s...

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Abstract

The invention provides an enzyme-linked immunosorbent assay kit capable of detecting dalapon in cotton and jute, and relates to the immunoassay detection technology. The kit includes an ELISA plate coated with dalapon, a dalapon antibody, a dalapon standard, a standard diluent, an enzyme-labelled secondary antibody, a substrate developing liquid, a washing buffer liquid and a stopping liquid. The key technology comprises that a monoclonal antibody which can identify dalapon is prepared. The kit adopts the high-specificity dalapon monoclonal antibody, a main reagent is provided in a form of a working liquid, the operation steps can be reduced, and the time is saved.

Description

technical field [0001] The invention belongs to the technical field of pesticide residue analysis and immunoassay detection, and in particular relates to the preparation of a monoclonal antibody capable of recognizing palapaquat and an enzyme-linked immunoassay kit. The invention discloses a preparation method of a specific monoclonal antibody, a coating agent and an immunogen and an enzyme-linked immunoassay method. Compared with the prior art, the monoclonal antibody prepared by the invention can recognize palapaquat, and the kit and method of the invention have the advantages of simplicity, speed, sensitivity, accuracy and the like. Background technique [0002] As a selective systemic herbicide with low toxicity and carcinogenicity, mutagenicity and teratogenicity, palapaquat is highly valued for its safety. [0003] At present, HPLC is the main method for the determination of thatraquat. Due to the shortcomings of this method, such as long analysis time, large sample v...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/577G01N33/531
Inventor 杜道林戴蔚蔚洪霞
Owner JIANGSU WISE SCI & TECH DEV
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