Efficient protein-to-membrane transferring buffer solution and preparation method thereof

A buffer and membrane transfer technology, which is applied in the field of high-efficiency protein transfer membrane buffer and its preparation, can solve the problems of PAGE gel and solid support displacement, low protein transfer effect, and increased temperature of electrophoretic solution, so as to shorten time, Improve the film effect and simplify the operation

Inactive Publication Date: 2016-08-17
SUZHOU NEW CELL & MOLECULAR BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

When using this transfer buffer system to transfer proteins, the temperature of the electrophoresis solution will increase significantly with time, high temperature will reduce the transfer effect, and will also cause displacement of PAGE gel and solid support, deformation of protein bands, etc. , so it is often necessary ...

Method used

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  • Efficient protein-to-membrane transferring buffer solution and preparation method thereof
  • Efficient protein-to-membrane transferring buffer solution and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] (1) Weigh the following reagents and place them in a 1000ml beaker:

[0031] Tris 4g; Taurine 0.07mol; SDS 0.15g;

[0032] (2) Add about 600ml of deionized water to the beaker, stir well;

[0033] (3) Add 50-100ml of methanol, add deionized water to dilute the solution to 1000ml and store at room temperature.

Embodiment 2

[0035] (1) Weigh the following reagents and place them in a 1000ml beaker:

[0036] Tris 0.01mol; Taurine 0.01mol; SDS 0.1g;

[0037] (2) Add about 600ml of deionized water to the beaker, stir well;

[0038] (3) Add 50-100ml of methanol, add deionized water to dilute the solution to 1000ml and store at room temperature.

Embodiment 3

[0040] (1) Weigh the following reagents and place them in a 1000ml beaker:

[0041] Tris0.1mol; Taurine 0.2mol; SDS 0.5g;

[0042] (2) Add about 600ml of deionized water to the beaker, stir well;

[0043] (3) Add 50-100ml of methanol, add deionized water to dilute the solution to 1000ml, and store it at room temperature.

[0044] (1) Use the transfer buffer prepared in Examples 1-3 to detect the effect on the transfer of PAGE gel to PVDF membrane, take 5ul of PageRuler TM Prestained Protein Ladder samples were added to the PAGE gel wells, and after the electrophoresis was completed, the membrane transfer experiment was performed according to the above method. The membrane transfer condition is 0.3A constant current, the time is 20 minutes, stop the electrophoresis, take out the sample and observe it, if figure 1 As shown, proteins with different molecular weights were completely transferred to the PVDF membrane, and no protein residues were observed on the PAGE gel.

[00...

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Abstract

The invention discloses a high-efficiency protein transfer membrane buffer, which comprises the following components: the concentration of Tris is 10mM-100mM; the concentration of taurine is 10mM-200mM. Compared with the traditional membrane transfer buffer commonly used at present, the high-efficiency membrane transfer buffer of the present invention uses a taurine component with a stronger buffer capacity, which improves the buffer capacity of the buffer during electrophoresis, and is suitable for proteins with a wide range of molecular weights. Molecules have high membrane transfer efficiency, which can shorten the time for protein transfer to the membrane. At the same time, the heat generated by the buffer during electrophoresis is low, and there is no need to use ice packs to cool down, which simplifies the operation during membrane transfer. The concentration of methanol is lower, and a better transfer effect can be achieved at a concentration of 10%.

Description

technical field [0001] The invention relates to a high-efficiency protein transmembrane buffer and a preparation method thereof. Background technique [0002] Western blotting, Western Blot, is a common experimental method in molecular biology, immunogenetics and biochemistry. It is to transfer the electrophoresis-separated cells or tissue proteins from the gel to a solid support, such as nitrocellulose. A protein detection technology that uses plain film (NC film), polyvinylidene fluoride film (PVDF film), and specific antibodies to detect specific antigens. Now it has been widely used in antibody activity detection, early disease diagnosis and protein level expression research and other aspects. Membrane transfer is an important step in Western blotting experiments. Proteins separated by polyacrylamide gel (PAGE) electrophoresis need to go through the membrane transfer step, that is, transfer from the PAGE gel to the solid phase support, before they can be processed by va...

Claims

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Application Information

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IPC IPC(8): G01N27/447
CPCG01N27/447
Inventor 杨勇刘永双
Owner SUZHOU NEW CELL & MOLECULAR BIOTECH CO LTD
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