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Pre-prepared colloid

A technology of prefabricated gel and separating gel, which is applied in the direction of biological testing, material inspection products, etc., can solve the problems of electrophoretic strip diffusion, short storage time, time consumption and manpower, etc., to increase flexibility and uniformity, storage and use Convenience, increased stability effect

Inactive Publication Date: 2003-01-15
GENETECH BIOTECH SHANGHAI
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Problems solved by technology

[0005] During electrophoresis, due to the high pH value of the buffer system, the conformation of the protein may change due to the easy oxidation of cysteine ​​to form a disulfide bond, resulting in a decrease in the separation efficiency of macromolecules; and proteins with different conformations are mixed together , will cause the diffusion of the electrophoretic band
In addition, acrylamide is also easy to hydrolyze, which greatly reduces the storage time and effect of the colloid; and the amino group of the protein is easy to react quickly with unpolymerized acrylamide, which may affect the size analysis of the protein
In addition, in order to ensure the experimental effect, most laboratories use freshly prepared glue, or the storage time is very short, which affects the efficiency and consumes a lot of time and manpower

Method used

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Embodiment Construction

[0031] Preparation of stock solutions: Except for TEMED from Sigma Chemical Co. and ammonium persulfate from Fluka Biochemika, the rest of the reagents were from Shanghai Sangon Bioengineering Co., Ltd. subpackaged products. in:

[0032]7×Bis-Tris (2.5M, pH6.5): Dissolve 26g of Bis-Tris in 1.5M HCl solution, adjust the pH to 6.5, and finally adjust the volume to 50ml.

[0033] 30% Acrylamide: Dissolve 29g of acrylamide and 1g of bis-methylene acrylamide in 100ml of distilled water.

[0034] 10% ammonium persulfate: 100 mg of ammonium persulfate was dissolved in 1 ml of distilled water.

[0035] 20×MOPS-SDS electrophoresis buffer: 104.6g MOPS / 60.6g Tris Base / 10g SDS / 3g EDTA dissolved in 500ml distilled water, the final pH should be 7.7.

[0036] All the above solutions were filtered with a 0.45 μm filter membrane and stored at 4°C.

[0037] The glue making process taking the prefabricated glue with a concentration of 12% as an example: according to image 3 Install the glas...

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Abstract

A prefabricated colloid used for electrophoretic analysis of protein contains polyacrylamide gel as main component, storage liquid (pH 6.5), neutral Bis-Tris buffering system (pH 6.5), ammonium persulfate, neutral MOPS-SDS electrophorectic buffering liquid and EDTA. Its advantages are high stability and easy storage and application.

Description

technical field [0001] The precast gel of the invention relates to a precast gel system used in protein electrophoresis analysis and a method for using the precast gel. Background technique [0002] Electrophoresis can be used to separate complex protein mixtures, study protein subunit composition and confirm protein homogeneity, etc., and can also purify proteins for subsequent research work. Almost all protein electrophoretic analyzes are performed on polyacrylamide gels, and the conditions used are always to ensure the dissociation of proteins into individual polypeptide subunits and minimize their aggregation with each other. The most commonly used method is to use the strong anionic detergent SDS together with a certain reducing agent, and dissociate the protein by heating, and then sample it on the electrophoresis gel. Denatured polypeptides bind to SDS and are therefore negatively charged. Since the amount of SDS bound to a polypeptide is almost always proportional t...

Claims

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Application Information

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IPC IPC(8): G01N33/68
Inventor 张涛李宾彭永济钱静李智任一萍
Owner GENETECH BIOTECH SHANGHAI
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