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Regeneration solution for Western blot membrane

An immunoblotting and protein technology, applied in biological testing, material testing products, measuring devices, etc., can solve the problems of undetectable, time-consuming and troublesome, and changes in the state of imprinted membrane proteins.

Active Publication Date: 2017-10-20
北京爱普拜生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In the usual western blot detection process, the blotted membrane loaded with protein can only be tested once at a time, and the internal reference protein cannot be detected on the same membrane at the same time. If the internal reference protein is to be detected, it can only be performed again The same series of protein immunodetection operations such as protein electrophoresis transfer to membranes are not only time-consuming and laborious, but also lead to changes in the state of proteins on the blotted membrane caused by repeated loading and operations.

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  • Regeneration solution for Western blot membrane
  • Regeneration solution for Western blot membrane
  • Regeneration solution for Western blot membrane

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0086] Example 1 Using high-efficiency western blot membrane regeneration solution to treat PVDF membrane and combine with chemiluminescence to detect the results of western blot membrane

[0087] 1. Experimental method

[0088] (1) Mouse liver tissue was added to 100 mg of animal tissue in a proportion of 1 ml by adding a general-purpose high-efficiency total protein extraction reagent and homogenized;

[0089] (2) Incubate on ice for 45 minutes;

[0090] (3) Centrifuge at 13,000 rpm for 30 minutes at 4°C;

[0091] (4) The collected supernatant is the total animal protein extract, which can be used for subsequent experimental operations.

[0092] (5) Take 24 μL of the extracted protein solution, add 6 μL of 5×loading buffer and mix thoroughly, heat and boil at 100°C for 5 minutes, place it at room temperature and then 4°C, centrifuge at 13000rpm for 20 minutes and absorb the supernatant for later use.

[0093] (6) Determination of protein concentration: the determination o...

Embodiment 2

[0115] Example 2 Using high-efficiency western blot membrane regeneration solution to treat PVDF membrane and combine with substrate chromogenic method to detect the results of western blot membrane

[0116] 1. Experimental method

[0117] (1) Mouse liver tissue was added to 100 mg of animal tissue in a proportion of 1 ml by adding a general-purpose high-efficiency total protein extraction reagent and homogenized;

[0118] (2) Incubate on ice for 45 minutes;

[0119] (3) Centrifuge at 13,000 rpm for 30 minutes at 4°C;

[0120] (4) The collected supernatant is the total animal protein extract, which can be used for subsequent experimental operations.

[0121] (5) Take 24 μL of the extracted protein solution, add 6 μL of 5×loading buffer, mix thoroughly, heat and boil at 100°C for 5 minutes, place it at room temperature and then 4°C, centrifuge at 13000rpm for 20 minutes and absorb the supernatant for later use.

[0122] (6) Determination of protein concentration: the determi...

Embodiment 3

[0144] Example 3 Using high-efficiency western blot membrane regeneration solution to treat NC membrane and combine with chemiluminescence to detect the results of western blot membrane

[0145] experimental method

[0146] (1) Mouse liver tissue was added to 100 mg of animal tissue in a proportion of 1 ml by adding a general-purpose high-efficiency total protein extraction reagent and homogenized;

[0147] (2) Incubate on ice for 45 minutes;

[0148] (3) Centrifuge at 13,000 rpm for 30 minutes at 4°C;

[0149] (4) The collected supernatant is the total animal protein extract, which can be used for subsequent experimental operations.

[0150] (5) Take 24 μL of the extracted protein solution, add 6 μL of 5×loading buffer, mix thoroughly, heat and boil at 100°C for 5 minutes, place it at room temperature and then 4°C, centrifuge at 13000rpm for 20 minutes and absorb the supernatant for later use.

[0151] (6) Determination of protein concentration: the determination of protei...

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Abstract

The invention belongs to the field of molecular biology and mainly relates to a regeneration solution for a Western blot membrane. With the adoption of the regeneration solution for the Western blot membrane, primary antibodies and secondary antibodies on the Western blot membrane can be effectively removed, so that repeated efficient utilization of the blotting membrane with transferred protein during Western blot detection is facilitated, the used blotting membrane can be quite conveniently reused for detecting other target proteins, time and labor are saved, furthermore, errors caused by re-sampling of the proteins can be eliminated, and comparability is enhanced. Detection experiments of multiple antibodies prove that by the aid of the regeneration solution for the Western blot membrane, the western blot membrane can be reused at least three times, so that correlative charges consumed by Western blot detection of conventional molecular biology experiments are greatly saved, and reuse of the Western blot membrane can be realized in about 30 min with the reagent. The regeneration solution is compatible with a substrate color-developing method, a chemiluminiscence color-developing method and a direct inspection method of fluorescence labeled secondary antibodies.

Description

technical field [0001] The invention relates to a protein immunoblotting membrane regeneration solution, which can quickly, efficiently and fully remove the primary antibody and secondary antibody bound on the protein immunoblotting membrane, so that the protein immunoblotting membrane can be used repeatedly. Background technique [0002] Proteomics refers to taking all proteins encoded by the genome as the research object, studying the composition and changing rules of proteins from the overall level of cells or tissues, so as to deeply understand the various physiology and pathology of organisms. Compared with traditional protein research, proteomics research embodies the characteristics of comprehensiveness, integrity, high throughput and large scale. The study of proteomics plays an irreplaceable important role in the empirical analysis of the theoretically predicted proteome that has completed the genome project. Proteomics technology is relatively complex, including t...

Claims

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Application Information

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IPC IPC(8): G01N33/53
CPCG01N33/5306
Inventor 于祥春冯晓燕林挺王文利龚建
Owner 北京爱普拜生物技术有限公司
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