Sudan 1 capable of being directly marked and preparation method and application thereof

A Sudan red and labeling technology, applied in the field of immunological detection, can solve the problem of no amino group or carboxyl group, etc., and achieve the effect of simple and easy method, high efficiency and simple operation.

Inactive Publication Date: 2011-07-06
NANJING UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, Sudan Red I itself does not have amino or carboxyl groups that can be directly coupled to carrier proteins, so it is necessary to introduce

Method used

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  • Sudan 1 capable of being directly marked and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Example 1 Synthesis of Directly Labelable Sudan Red I

[0024] 1. Dissolve p-phenylenediamine (2.0g, 18.5mmol) in 30ml of anhydrous tetrahydrofuran, under stirring conditions, dissolve acetic anhydride (1.7ml, 18.5mmol) in 4ml of anhydrous tetrahydrofuran within 1 hour Add dropwise to the p-phenylenediamine solution, TLC monitors whether the reaction is complete;

[0025] 2. Filter the reaction solution, leave the precipitate, and then recrystallize with methanol to obtain the monoamino group protected product;

[0026] 3. Dissolve the monoamino-protected sample (0.5g, 3.3mmol) in 50ml of ice water containing 2.5ml of concentrated HCl; add sodium nitrite (0.228g, 3.3mmol) dissolved in 1ml of ice water to the above solution slowly After the dropping, continue to stir and react for 15 minutes to obtain the diazonium salt solution;

[0027] 4. Add β-naphthol (0.488g, 3.3mmol) dissolved in 2ml 10% NaOH solution dropwise to the above diazonium salt solution. After the addition is...

Embodiment 2

[0029] Example 2 Directly labelable biotin labeling of Sudan Red I

[0030] First, acylate biotin with acid chloride, then add it to the anhydrous tetrahydrofuran solution of Sudan Red I that can be directly modified, and add a small amount of pyridine to the reaction system to absorb the hydrogen chloride released by the reaction. After the reaction is completed, the biotin-labeled Sudan Red I conjugate is obtained by column purification. On the one hand, this conjugate can competitively bind to Sudan Red I monomer; on the other hand, because 4 biotins can interact with one streptavidin Harmonic molecules are combined, so when used in a competitive one-step detection system, the sensitivity of detection can be improved.

Embodiment 3

[0031] Example 3 Enzyme labeling of Sudan Red I that can be directly labeled

[0032] At present, most of the enzymes used in the immunoassay system for small molecule substances are horseradish peroxidase (HRP) and alkaline phosphatase (AP). In many existing competitive one-step detection systems, small The coupling ratio of molecule and enzyme is mostly 1:1, because these small molecules are mainly dehydrated with amino groups on enzyme molecules to form amides. For enzymes, the number of amino groups available for coupling in their structure is significantly lower than Number of carboxyl groups. According to reports, for horseradish peroxidase, only one amino group can be used for coupling. This easily leads to the small molecule substance itself being too small and too small to be encapsulated in the enzyme molecule, thereby reducing the sensitivity of detection. Therefore, we use the amino-modified Sudan Red I to incorporate multiple small molecules into the enzyme molecu...

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Abstract

The invention belongs to the technical field of immunological test, and in particular relates to a method for preparing Sudan 1 capable of being directly marked and application of the Sudan 1. In the method, a technical route of denovo synthesis is mainly utilized, amino active group is introduced successfully on the premise of not changing an antigenic determinant in a host molecular structure of the Sudan 1, and a trim capable of directly being connected with fluorescent substances and enzymes is provided for the Sudan 1; and an important problem of competitive antigen preparation is solved for establishing a Sudan 1 competitive method enzyme-linked immuno sorbent assay (ELISA) detection system. The whole method is simple and practicable, easy to operate, and high in efficiency of connecting the fluorescent substances and the enzymes.

Description

technical field [0001] The invention belongs to the technical field of immunological detection, and in particular relates to a preparation method and application of Sudan Red I which can be directly labeled. Background technique [0002] Sudan Red I is a synthetic azo dye, which is insoluble in water and has good lipophilicity. As a commonly used industrial dye, it is widely used in the color enhancement of solvents and the enhancement of textiles. [0003] Since the incidents related to the contamination of food by Sudan Red I were exposed in the early 21st century, people have paid more and more attention to the harm caused by Sudan Red I. Studies have shown that the toxicity of Sudan Red I is closely related to the amines and naphthols produced by its metabolism. When people eat or touch food containing Sudan Red I, it can enter the body through the human skin or digestive system. The liver is the main target organ for the carcinogenicity of Sudan Red I. On the one han...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07C245/10G01N33/532
Inventor 沈萍萍徐加发范祚舟
Owner NANJING UNIV
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