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Novel Clear Native electrophoresis method utilizing aromatic sulfonic acid compound

A compound and electrophoresis technology, applied in the fields of peptide preparation, chemical instruments and methods, organic chemistry, etc., can solve the problem of the absence of multimeric proteins

Inactive Publication Date: 2011-11-02
JAPAN SCI & TECH CORP +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, it has the following problem: for a certain protein, a multimeric protein that does not exist in the natural state is formed

Method used

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  • Novel Clear Native electrophoresis method utilizing aromatic sulfonic acid compound
  • Novel Clear Native electrophoresis method utilizing aromatic sulfonic acid compound
  • Novel Clear Native electrophoresis method utilizing aromatic sulfonic acid compound

Examples

Experimental program
Comparison scheme
Effect test

preparation example Construction

[0182] Samples can be prepared by known methods. For example, a sample can be prepared by mixing the above protein sample in a buffer composition containing compound (I) and a sample buffer. In addition, a sample can be prepared by adding a reagent for electrophoresis containing compound (I) to a buffer obtained by solubilizing a protein sample, and then adding other components such as glycerin as necessary.

[0183] The protein concentration in the sample varies according to the staining method or detection method after electrophoresis, and cannot be set to a fixed value. When detecting by CBB staining, it is usually about 0.5 μg / well to about 50 μg / well, preferably about 50 μg / well. 1 μg / well to about 10 μg / well. In addition, when preparing a fluorescent protein fusion product and detecting the fluorescence of the protein, it is generally 0.05 μg / well to 1 μg / well, preferably about 0.1 μg / well to about 0.5 μg / well. In addition, when performing silver staining and detection...

Embodiment 1

[0220] 1. Production of compounds represented by general formula (Ig)

[0221] At 0°C, in 12.0 g (14.1 mmol) of (Benzenemethanaminium), N-[4-[[4-[(4-ethoxyphenyl)amino]phenyl][4-[ Ethyl[(3-sulfophenyl)methyl]amino]phenyl]methylene]-2,5-cyclohexadiene-1-ylidene]-N-ethyl-3-sulfo-hydrogen Oxide, inner salt, monosodium salt) (CBB G250) were added methanol (60 ml) and 2.65 g of sodium cyanoborohydride (sodium cyanoborohydride) (42.3 mmol), and stirred at room temperature for 1.5 hours.

[0222] After the reaction solution was concentrated under reduced pressure, it was recrystallized by mixing and dissolving diethyl ether and methanol to obtain 8.95 g of 3,3'-(4,4'-((4-(4-ethoxybenzene Amino)phenyl)methylene)bis(3-methyl-4,1-phenylene))bis(ethylazanediyl)bis(methylene)dibenzenesulfonate ( Hereinafter, it is referred to as compound Ig) (yield 73%).

[0223] The NMR analysis results of the obtained compound (Ig) are shown below. 1 H NMR (270MHz, CD 3 OD): δ7.65-7.85(m, 4H), 7.37...

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Abstract

Disclosed is a reagent for the electrophoresis of a protein. Also disclosed is a gel or buffer composition for electrophoresis, which contains the reagent; and a protein separation method and an electrophoresis kit, each of which utilizes the reagent and the composition.

Description

technical field [0001] The present invention mainly relates to a reagent for electrophoresis, a composition for electrophoresis, a kit for electrophoresis and a protein separation method. Background technique [0002] Nowadays, Blue-Native electrophoresis is often used as a method to study the molecular weight and aggregation state of membrane proteins and supramolecular complexes while maintaining the natural state or enzyme activity. The above method was developed by Schagger et al. (refer to Non-Patent Document 1), and the negatively charged Coomassie Brilliant Blue (Coomassie Brilliant Blue, hereinafter referred to as "CBB") G250 utilizes its combination with proteins to bias the total charge of the protein towards negative charges, Then, a voltage is applied to proteins in an acrylamide gel having a fine network structure to separate them according to their molecular size. However, since the electrophoresis buffer has a blue color derived from CBB G250, it is inconveni...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N27/447C07C309/46
CPCC07C309/46G01N27/44747C07K1/26G01N27/447G01N33/68
Inventor 日野智也村田武士岩田想菅敏幸
Owner JAPAN SCI & TECH CORP
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