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Method for increasing mass spectrum identification number of protein and/or peptide fragment group by using polystyrene material

A polystyrene-based, identification method technology, applied in the field of proteomics, to achieve the effects of reducing human error, simple operation, and avoiding interference

Pending Publication Date: 2022-02-11
PROTEINT (TIANJIN) BIOTECHNOLOGY CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there are no reports on the use of such materials to adsorb proteins.

Method used

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  • Method for increasing mass spectrum identification number of protein and/or peptide fragment group by using polystyrene material
  • Method for increasing mass spectrum identification number of protein and/or peptide fragment group by using polystyrene material
  • Method for increasing mass spectrum identification number of protein and/or peptide fragment group by using polystyrene material

Examples

Experimental program
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Effect test

Embodiment 1

[0033] 1. Take out different plasma samples from the refrigerator, take 200 μL of plasma from each sample and thaw at 37°C, centrifuge at 1,500g for 10 minutes at 4°C, remove the bottom precipitate and transfer the supernatant to a new tube;

[0034] 2. Take 100 μL plasma and add 100 μL binding buffer (50 mM Tris, 10 mM EDTA) to resuspend polystyrene microspheres (particle size 1 μm, CAS number: 9003-53-6), and incubate at room temperature for 10-60 minutes;

[0035] 3. After the incubation is completed, centrifuge at 12,000g for 10 minutes to remove the supernatant, resuspend the pellet with 300 μL washing buffer reagent (50mM Tris, 10mM EDTA), incubate at room temperature for 5 minutes, and centrifuge at 12,000g for 5 minutes to remove the supernatant;

[0036] 4. Repeat step 3 twice;

[0037] 5. Add the precipitate obtained by combining the original plasma and materials into protein electrophoresis Loading Buffer and boil, then run electrophoresis on the supernatant, the re...

Embodiment 2

[0060] In order to prove that the use of this material can be used for the enrichment of micro samples and enzymatic desalination at the protein level or peptide level, it can be used to replace the existing traditional SP3 mixed magnetic bead materials. In this example, the SP3 mixed material was used as the control group, and the polystyrene material in Example 1 was used as the carrier to conduct proteomic analysis experiments on Hela cell protein and peptide extracts:

[0061] Take the Hela cell extract protein standard as an example:

[0062] 1. Take two 100 μg Hela cell protein samples dissolved in 40uL buffer (4% SDS, 100mM Tris, pH 8.0) for reductive alkylation;

[0063] 2. Add 10μL stored 10μg / μL SP3 mixed beads and the same amount of polystyrene microspheres (particle size between 50nm and 100μm, CAS number: 9003-53-6), then add 150uL buffer and 200uL acetonitrile As a binding buffer, shake and mix gently, and react at room temperature for 8 minutes;

[0064] 3. Af...

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Abstract

The invention discloses a method for improving the mass spectrum identification number of a protein and / or peptide fragment group by using a polystyrene material. The method comprises the following steps of: adding a binding buffer solution into a sample to be detected, and adding the polystyrene material to obtain a suspension; incubating the suspension, performing high-speed centrifugation, removing supernate, and retaining a precipitate I; re-suspending the precipitate I by using a cleaning buffer solution to obtain a re-suspension, centrifuging the re-suspension at a high speed, removing a supernatant, retaining a precipitate II, and repeating the step for several times; and preparing the precipitate II into a proteome / peptide fragment group sample, and carrying out mass spectrometric detection. The polystyrene material and the corresponding buffer solutions are adopted, different kinds of serum / plasma proteins and peptide fragments can be effectively adsorbed, selective enrichment of low-abundance proteins and peptide fragments in serum / plasma is achieved, interference of high-abundance proteins in the serum / plasma on identification of the low-abundance proteins during mass spectrum identification is effectively avoided. The method can be used for identifying various protein spectrums.

Description

technical field [0001] The invention belongs to the technical field of proteomics, and in particular relates to a method for increasing the mass spectrum identification number of proteins and or peptide groups by using polystyrene materials. Background technique [0002] Since serum / plasma is an important part of blood, which contains substances released by various tissues and organs of organisms, there is a large amount of physiological or pathological information that can be discovered. It is speculated that there are more than 10,000 kinds of proteins in serum / plasma, but due to the limited detection and identification technology at present, only a small part of the proteins can be detected. Since serum / plasma contains dozens of high-abundance proteins, which account for more than 95% of the total protein content of serum / plasma, it brings great challenges to the detection and separation of plasma / serum proteins. At present, without removing high-abundance proteins, only...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/02G01N30/72
CPCG01N30/02G01N30/72
Inventor 马聪聪李嘉欣赵瑾原续波陈亮宇李捷宋雷任丽
Owner PROTEINT (TIANJIN) BIOTECHNOLOGY CO LTD
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