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Beta-amyloid 1-42 oligomer as well as preparation method and identification method therefor

An oligomer and tetramer technology, applied in the field of identification, Aβ1-42 oligomer and its preparation, can solve the problems of complex composition, unstable oligomer, low yield and the like, and achieve single composition and preparation method. Simple, high-yield effects

Inactive Publication Date: 2015-10-14
广西南宁灵康赛诺科生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Aiming at the deficiencies of the existing preparation and synthesis of Aβ1-42 oligomers, such as instability, complex components, and low yield, the present invention provides a newly synthesized Aβ1-42 oligomer

Method used

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  • Beta-amyloid 1-42 oligomer as well as preparation method and identification method therefor
  • Beta-amyloid 1-42 oligomer as well as preparation method and identification method therefor
  • Beta-amyloid 1-42 oligomer as well as preparation method and identification method therefor

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0063] A method for preparing Aβ1-42 oligomers, comprising the following steps:

[0064] 1. Add 1 mg of Aβ1-42 synthetic peptide to 220 μL HFIP, and let it stand at room temperature for 30 minutes to obtain a completely dissolved solution;

[0065] 2. Transfer the solution obtained in step 1 to a siliconized centrifuge tube, and blow it on a dry bath nitrogen blower for 8 minutes to completely dry the HFIP, and obtain a transparent sheet-like solid that sinks to the bottom of the tube;

[0066] 3. Add 40 μl DMSO to the centrifuge tube to dissolve the solid at the bottom of the centrifuge tube completely, then add a 10 mM HEPES buffer solution with a pH value of 7.4 to make the final final volume of the solution 1 ml, mix well, and get final product.

Embodiment 2

[0068] A method for preparing Aβ1-42 oligomers, comprising the following steps:

[0069] 1. Add 1 mg of Aβ1-42 synthetic peptide to 225 μL HFIP, and let stand at room temperature for 60 minutes to obtain a completely dissolved solution;

[0070] 2. Transfer the solution obtained in step 1 to a siliconized centrifuge tube, and blow it on a dry bath nitrogen blower for 15 minutes to completely dry the HFIP, and obtain a transparent sheet-like solid that sinks to the bottom of the tube;

[0071] 3. Add 40 μl DMSO to the centrifuge tube to dissolve the solid at the bottom of the centrifuge tube completely, then add a 10 mM HEPES buffer solution with a pH value of 7.4 to make the final final volume of the solution 1 ml, mix well, and get final product.

[0072] 3. Identification method of Aβ1-42 oligomer

Embodiment 3

[0074] A method for identifying Aβ1-42 oligomers, comprising the following steps:

[0075] 1. SDS-polyacrylamide gel electrophoresis: Take 2 μL of the Aβ1-42 oligomer solution obtained in Example 1, mix it with 18 μL 4X electrophoresis sample buffer, and then load the sample. The electrophoresis sample volume is 5ul / swimming lane. The gel used for electrophoresis is 4–12% gradient gel, and 10 μL of pre-stained molecular weight standard with a molecular weight of 3.6-260KD is added to one lane, and the voltage of 80V is selected for electrophoresis for 30 minutes, and then the voltage of 100v is used to continue electrophoresis for 2 hours;

[0076] 2. Membrane transfer: transfer the electrophoresis protein band to the PVDF membrane, transfer the membrane by semi-dry method, the time is 6 minutes, cut the transferred PVDF membrane according to the size of the sample lane, and then soak the cut PVDF membrane into the TBST solution for 5 minutes;

[0077] 3. Sealing: Soak the PV...

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Abstract

The invention discloses a beta-amyloid 1-42 oligomer. The beta-amyloid 1-42 oligomer is a mixture of oligomers and monomers mainly; the oligomers are identified through a Western blot method: the molecular weight is 12-20 KD, and the oligomers are tripolymers and tetramers; and the monomers are identified through the Western blot method: the molecular weight is 4-4.5 KD. The invention also discloses a preparation method and an identification method of the beta-amyloid 1-42 oligomer. The beta-amyloid 1-42 oligomer provided by the invention is relatively single in composition and relatively stable in product, and a good foundation is provided for establishing ideal and reliable AD models; the preparation method is simple, high in yield and time-saving; and the adopted SDS-PAGE in combination with the Western blot method has the advantages of high sensitivity, high specificity, convenience, high speed, simple operation, relatively low cost and the like.

Description

technical field [0001] The invention relates to the field of beta-amyloid protein, in particular to an Aβ1-42 oligomer and its preparation and identification method. Background technique [0002] Alzheimer's disease (Alzheimer's disease, AD), also known as senile dementia, is a chronic, persistent, degenerative brain degenerative disease that occurs in old age or early old age, and its clinical manifestations are memory loss, cognition disorders, personality changes, etc. The pathological features of the disease mainly include extracellular senile plaques with β-amyloid (Aβ) as the core and intracellular neurofibrillary tangles in the cerebral cortex and hippocampus. In the United States, AD is the fourth leading cause of death after cardiovascular disease, cancer and stroke. According to statistics, the number of AD patients in the world is as high as 25 million, and the number of AD patients in my country has reached more than 6 million, ranking first in the world. The du...

Claims

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Application Information

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IPC IPC(8): C07K14/47G01N33/68
CPCC07K14/4711G01N33/6896G01N2333/4709
Inventor 岳峰陆春玲
Owner 广西南宁灵康赛诺科生物科技有限公司
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