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Application of dansylhydrazine and derivatives thereof in specific detection of glycoprotein

A technology of dansylhydrazide and its derivatives, which is applied in the field of application of dansylhydrazine and its derivatives in the specific fluorescence detection of glycoproteins, can solve the problem of inability to achieve economic benefits and experimental development together, unfavorable development of basic biotechnology research, The cost of biological experiments is not high, and the effects of low cost, simple and fast operation, and low environmental pollution are achieved.

Inactive Publication Date: 2015-10-14
WENZHOU MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, kits in the field of glycoprotein detection are mainly concentrated in foreign biological reagent companies, and most of the products are sold in the market at high value-added prices. The high price is not conducive to the development of basic biotechnology research.
In particular, the cost of using existing biological reagents to carry out biological experiments in China remains high, and it is impossible to achieve the common development of economic benefits and experiments

Method used

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  • Application of dansylhydrazine and derivatives thereof in specific detection of glycoprotein
  • Application of dansylhydrazine and derivatives thereof in specific detection of glycoprotein
  • Application of dansylhydrazine and derivatives thereof in specific detection of glycoprotein

Examples

Experimental program
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Effect test

Embodiment 1

[0029] Example 1 Dansylhydrazide glycoprotein-specific fluorescent staining

[0030] figure 1 is the chemical structural formula of dansyl hydrazide.

[0031] The dansylhydrazide glycoprotein fluorescent staining method is carried out as follows:

[0032] 1) Put the gel of the protein sample after SDS-PAGE electrophoresis in 40% ethanol and 10% acetic acid aqueous solution for 30 minutes, and discard the fixative;

[0033] 2) Oxidize in periodic acid solution for 20min, wherein the periodic acid solution is an aqueous solution of acetic acid containing 0.5% by weight and 3% by volume of periodic acid. Then rinse with 3% acetic acid aqueous solution by volume for 3 times, 5min each time;

[0034] 3) adding dyeing solution for dyeing for 30min, wherein the dyeing solution is dansyl hydrazide containing 0.006% by weight and 3% by volume of acetic acid aqueous solution;

[0035] 4) Add an eluent containing 3% acetic acid aqueous solution by volume, and elute twice for 10 minut...

experiment example 1

[0038] Experimental Example 1 The dansyl hydrazide glycoprotein fluorescent staining method was compared with other staining methods for the detection of standard proteins.

[0039] (A) Dansylhydrazide glycoprotein fluorescent staining method, (B) Pro-Q Emerald glycoprotein fluorescent staining method, (C) SYPRO Ruby whole protein fluorescent staining method. Pro-Q Emerald glycoprotein fluorescent staining method and SYPRO Ruby whole protein fluorescent staining method were both recorded in the literature; 8 different standard proteins from Sigma were used as samples. With 1,1000ng; with 2,500ng; with 3, 250ng; with 4, 125ng; with 5, 64ng; with 6, 32ng; with 7, 16ng; with 8, 8ng; with 9, 4ng; The result is as figure 2 As shown, it shows that the detection sensitivity of dansyl hydrazide glycoprotein fluorescent staining method is close to that of Pro-Q Emerald488 glycoprotein fluorescent staining method.

experiment example 2

[0040] Experimental Example 2 The dansyl hydrazide glycoprotein fluorescent staining method was compared with other staining methods for the detection of total protein in human serum.

[0041](A) Dansylhydrazide glycoprotein fluorescent staining method, (B) Pro-Q Emerald glycoprotein fluorescent staining method, (C) SYPRO Ruby whole protein fluorescent staining method. Pro-Q Emerald glycoprotein fluorescent staining method and SYPRO Ruby total protein fluorescent staining method were operated according to the literature; the extracted human serum total protein was used as the sample. With 1, 5000ng; With 2, 2500ng; With 3, 1250ng; With 4, 625ng; With 5, 312ng; With 6, 160ng; With 7, 80ng; With 8, 40ng; The result is as image 3 As shown, it shows that the detection sensitivity of dansyl hydrazide glycoprotein fluorescent staining method is close to that of Pro-Q Emerald488 glycoprotein fluorescent staining method.

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Abstract

The invention relates to a specific fluorescence detection technology of glycoprotein and particularly relates to an application of dansylhydrazine and derivatives thereof in specific detection of the glycoprotein. The invention also provides a method of performing specific fluorescence detection of the glycoprotein with the dansylhydrazine, wherein the method includes the steps of fixing a gel containing a protein sample after electrophoresis in a fixing liquid, oxidizing the gel with a periodic acid solution, washing the gel with acetic acid water solution; dyeing the gel, and eluting the gel. The technology is high in sensitivity and selectivity, is simple and rapid in operation, is good in repeatability, linear relationship and mass spectrum compatibility, is safe in use and is low in cost. The technology can be used for researching of high-flux proteomics well.

Description

technical field [0001] The invention relates to a glycoprotein-specific fluorescence detection technology, in particular to the application of dansyl hydrazide and its derivatives in glycoprotein-specific fluorescence detection. Background technique [0002] Protein glycosylation plays a crucial role in the field of life science research. As one of the most important and common protein post-translational modifications, it is currently known that at least half of mammalian proteins are glycosylated. Glycoproteins are widely distributed in tissues, cells, and body fluids, especially in the surface of cell membranes and body fluids. Conduction, cell attachment and many other important biological processes. Changes in the number of glycoproteins or changes in the structure of sugar chains may lead to disease. Not only are the diagnostic markers of many diseases known currently are glycoproteins, but also glycoproteins account for a high proportion of drugs certified by interna...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/64
Inventor 丛维涛朱忠欣玄元虎周旋
Owner WENZHOU MEDICAL UNIV
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