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SDS-PAGE (sodium dodecyl sulfate polyacrylamide gel electrophoresis) coomassie brilliant blue R250 fast staining solution, staining method and application

A Coomassie brilliant blue, rapid dyeing technology, applied in the preparation of test samples, material inspection products, measuring devices, etc., can solve the problems of increasing the probability of sample contamination, prolonging the operation time, cumbersome fixing and sensitization steps, etc. The effect of shortening the dyeing process time, background uniformity, and shortening of the dyeing time

Inactive Publication Date: 2012-07-25
INST OF TROPICAL BIOSCI & BIOTECH CHINESE ACADEMY OF TROPICAL AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the steps of fixation and sensitization before staining are cumbersome, which prolongs the operation time and increases the probability of sample contamination

Method used

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  • SDS-PAGE (sodium dodecyl sulfate polyacrylamide gel electrophoresis) coomassie brilliant blue R250 fast staining solution, staining method and application
  • SDS-PAGE (sodium dodecyl sulfate polyacrylamide gel electrophoresis) coomassie brilliant blue R250 fast staining solution, staining method and application
  • SDS-PAGE (sodium dodecyl sulfate polyacrylamide gel electrophoresis) coomassie brilliant blue R250 fast staining solution, staining method and application

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] (1), the rapid staining liquid that the present embodiment adopts:

[0035] Components containing the following contents: 30% ethanol by volume, 10% methanol by volume, CBB R250 with a mass concentration of 1.25g / L, 5% phosphoric acid by volume, and It is 100g / L ammonium sulfate.

[0036] Concretely formulated in the following ways:

[0037] Staining solution 11: Measure 300mL ethanol and 100mL methanol in a graduated cylinder, weigh 1.25g CBB R250 powder with an electronic balance, and dissolve it in pure water to 500mL, mix well and seal it, and place it overnight at 37°C at 100 rpm. The dye is fully dissolved.

[0038] Staining Solution 21 Weigh 100g of ammonium sulfate with an electronic balance, add 300mL of pure water to fully dissolve it, measure 59mL of 85% phosphoric acid in a graduated cylinder, and dilute to 500mL with pure water.

[0039] Fully mix the staining solution 11 and the staining solution 21, and name it as the staining solution R for later use. ...

Embodiment 2

[0056] (1), the rapid staining liquid that the present embodiment adopts:

[0057] Components containing the following contents: 30% ethanol by volume, 10% methanol by volume, CBB R250 with a mass concentration of 1.25g / L, 5% phosphoric acid by volume, and It is 100g / L ammonium sulfate.

[0058] Its preparation process is with embodiment 1.

[0059] (2), Extraction of Hevea brasiliensis C whey protein:

[0060] Using the BPP method to extract the C whey protein of Hevea brasiliensis (extraction method is the same as above), the collected fresh latex is subjected to ultracentrifugation to separate the C whey component, which is dissolved in an equal volume of protein lysate BPP, and the protein is extracted using Tris balanced phenol Extract, use supersaturated ammonium sulfate to precipitate protein, and lyse the protein precipitate to obtain protein solution. The Bradford method was used for protein quantification, and a unidirectional SDS-polyacrylamide gel electrophoresi...

Embodiment 3

[0076] (1), the component and the preparation method of the fast dyeing solution that the present embodiment adopts are the same as embodiment 1, and the dyeing solution R is evenly divided into two equal parts;

[0077] (2), cassava leaf protein, Hevea brasiliensis C whey protein, one-way SDS-PAGE electrophoresis of bovine serum protein:

[0078] Use SDS-PAGE with a mass percentage of 12.5% ​​to prepare two separation gels with a thickness of 1.5 mm. After the gels are coagulated, prepare a SDS-PAGE stacking gel with a mass percentage of 4%, and use a 15-tooth comb to make 15 wells Stacking gels with a maximum loading volume of 100 µl. Add 30 micrograms of the protein solution to the gel loading hole according to the loading order of cassava leaf protein, Brasilia C whey protein, and bovine serum protein, and use Hoefer SE 600chroma one-dimensional gel electrophoresis instrument to set constant power parameters , for one-way SDS-PAGE electrophoresis.

[0079] (3), staining ...

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Abstract

The invention discloses SDS-PAGE (sodium dodecyl sulfate polyacrylamide gel electrophoresis) coomassie brilliant blue R250 fast staining solution, which includes from 20 to 40% of ethanol by volume percentage, from 5 to 10% of carbinol by volume percentage, from 0.5 to 1.25g / L of CBB (coomassie brilliant blue) R250 by mass concentration, from 2.5 to 5% of phosphoric acid by volume percentage and from 100 to 200g / L of ammonium sulfate by mass concentration. A fixing step and a sensitization step before staining are omitted, protein staining time can be shortened, and the possibility that protein samples are polluted is greatly reduced. The invention further discloses a method for staining SDS-PAGE gel by the aid of the staining solution and application of the staining solution in a protein two-dimensional gel electrophoresis and protein western blot qualitative diagnosis and / or quantification process.

Description

technical field [0001] The present invention relates to a Coomassie Brilliant Blue fast staining solution, a dyeing method and an application thereof, in particular to a SDS-PAGE (sodium dodecyl sulfate-polyacrylamide gel) Coomassie Brilliant Blue R250 fast staining solution, a dyeing method and an application . Background technique [0002] Proteomics research can reveal the laws of life by constructing a map of temporal and spatial differences in protein expression in cells. The SDS polyacrylamide gel electrophoresis separation technology of protein is the key technology of proteomics research, and the staining of the gel is the core technology to obtain the analyzable map. In protein analysis and detection, Coomassie brilliant blue staining, silver nitrate staining and fluorescent dye staining are three staining methods widely used in protein analysis and detection. The silver nitrate staining method is recognized as a gel staining method with high sensitivity, and the ...

Claims

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Application Information

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IPC IPC(8): G01N1/30G01N27/447G01N33/68
Inventor 王旭初王海燕郭安平王丹常丽丽
Owner INST OF TROPICAL BIOSCI & BIOTECH CHINESE ACADEMY OF TROPICAL AGRI SCI
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