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Enzyme-linked immunoassay kit for detecting phenylethanolamine A by direct competition method

An enzyme-linked immunosorbent reagent, phenylethanolamine technology, applied in the direction of measuring devices, instruments, scientific instruments, etc., to achieve the effect of strong specificity, low pretreatment requirements, and high sensitivity

Inactive Publication Date: 2012-07-04
于洪侠
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The ELISA method of phenylethanolamine A has not been reported yet

Method used

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  • Enzyme-linked immunoassay kit for detecting phenylethanolamine A by direct competition method
  • Enzyme-linked immunoassay kit for detecting phenylethanolamine A by direct competition method
  • Enzyme-linked immunoassay kit for detecting phenylethanolamine A by direct competition method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] Example 1 Synthesis of Immunogen and Preparation of Immune Serum

[0023] 1.1 Reagents and instruments

[0024] Phenylethanolamine A (gifted by Hangzhou Dean Technology Co., Ltd.), succinic anhydride (purchased from Beijing Yaobei Biotechnology Co., Ltd.), pyridine (Pyridine, AR.WM=79.10, content > 99.5%, developed by Tianjin Kemiou Chemical Reagent center), N, N-dimethylformamide (Dimethylformamide, DMF, produced by ACROSORGANICS, New Jersey, USA, purchased from Bailingwei Chemical Reagent Company), EDC (purchased from Bailingwei Chemical Reagent Company), bovine serum albumin (BSA), Ovalbumin (OVA), etc. (purchased from Beijing Yaobei Biotechnology Co., Ltd.), and other reagents were of analytical grade.

[0025] Double-beam ultraviolet-visible spectrophotometer (TU-1909, Beijing Puxi General Instrument Co., Ltd.), chromatography device (3057 portable recorder, Chongqing Chuanyi No. 4 Factory; SBS series numerical control drop counter, constant flow pump, automatic p...

Embodiment 2

[0037] Example 2 Establishment of immunoassay method

[0038] 2.1 ELISA square array method to determine the optimal reaction concentration

[0039] Coat the microtiter plate with 100 μl per well of phenylethanolamine A antibody at a serial concentration of 1000 μg / ml, 100 μg / ml, 10 μg / ml, 5 μg / ml, 1 μg / ml, and 0.25 μg / ml, coat overnight at 4°C, and wash for 3 Once, pat dry, block overnight at 4°C with 200 μl of blocking solution per well, wash 3 times, and pat dry. Add 50 ul of phenylethanolamine A horseradish peroxidase marker diluted from 1:100, react at room temperature for 45 minutes, wash three times, add 50 μl of substrate A solution and 50 μl of substrate B solution, and incubate at room temperature for 15 minutes in the dark, 50 μl of stop solution was used to terminate the reaction, and the A value (450nm) was detected by a microplate reader. Set two parallels at the same time, and take the coating concentration when the OD value is about 1.5 as the optimal concent...

Embodiment 3

[0047] Example 3 Establishment of an enzyme-linked immunosorbent assay kit for detecting phenylethanolamine A

[0048] An enzyme-linked immunosorbent assay kit for detecting phenylethanolamine A was set up to include the following components:

[0049] (1) a microtiter plate coated with phenylethanolamine A antibody;

[0050] (2) phenylethanolamine A horseradish peroxidase marker working solution;

[0051] (3) 6 bottles of phenylethanolamine A standard solution, the concentrations were 0 μg / L, 0.1 μg / L, 0.3 μg / L, 0.9 μg / L, 2.7 μg / L, 8.1 μg / L;

[0052] (4) Substrate chromogenic solution A is hydrogen peroxide or carbamide peroxide, and substrate chromogenic solution B is tetramethylbenzidine or OPD;

[0053] (5) The washing solution is a phosphate buffer containing 0.05% Tween 20;

[0054] (6) The concentrated sample diluent is a phosphate buffered saline solution of 0.05% Tween-20;

[0055] (7) The stop solution is 2mol / L hydrochloric acid solution.

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Abstract

The invention discloses an enzyme-linked immunoassay kit for detecting phenylethanolamine A by a direct competition method. The enzyme-linked immunoassay kit for detecting phenylethanolamine A provided by the invention comprises an enzyme label plate coated by phenylethanolamine A specific antibodies, and a phenylethanolamine A horseradish peroxidase marker. The enzyme-linked immunoassay kit for detecting phenylethanolamine A of the invention is sensitive, rapid, accurate, and is mainly used for screening of large quantities of samples; main reagents in the kit are provided in the form of operating fluid, and the kit is convenient for using, has the characteristics of high specificity, high sensitivity, high precision, high accuracy, and the like, can rapidly detect residual phenylethanolamine A in feed and animal products.

Description

technical field [0001] The invention relates to the fields of ELISA and veterinary drug residue detection. Specifically, the present invention relates to an enzyme-linked immunosorbent assay kit for detecting phenylethanolamine A. technical background [0002] Phenylethanolamine A, also known as clenrapmine CLRA, is a kind of artificial Synthetic chemicals. Clonpamine is an isomer of formoterol and a by-product of ractopamine synthesized by Eli Lilly and Company of the United States. It has the same action and effect as clenbuterol and ractopamine, and belongs to a type of β2-agonist . It is listed as a substance prohibited to be used in feed and animal drinking water by the Ministry of Agriculture Announcement No. 1519. Therefore, detection methods must be established to prohibit the occurrence of such incidents and ensure food safety. [0003] Most of the existing detection methods for phenylethanolamine A are chromatography, but the sensitivity of these methods is gr...

Claims

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Application Information

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IPC IPC(8): G01N33/543
Inventor 于洪侠
Owner 于洪侠
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