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Protein collection and enzymolysis integrated sample pretreatment apparatus and its production

A sample pretreatment and protein technology, applied in the preparation of test samples, etc., can solve the problems of only suitable enzyme column reactors, difficulty in matching flow rates, and decreased activity, and achieve low cost, fast enzymatic hydrolysis, and simple production methods Effect

Inactive Publication Date: 2009-07-01
DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

(1) The conditions of protein enzymatic hydrolysis are usually harsh, and it is difficult to be compatible with the optimal conditions for protein separation, especially the flow rate is difficult to match; (2) The activity of the enzyme column will decrease after processing a certain amount of sample, and it is difficult to regenerate; (3) Currently Enzyme column reactors are only suitable for processing microliter or nanoliter samples

Method used

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  • Protein collection and enzymolysis integrated sample pretreatment apparatus and its production
  • Protein collection and enzymolysis integrated sample pretreatment apparatus and its production
  • Protein collection and enzymolysis integrated sample pretreatment apparatus and its production

Examples

Experimental program
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Effect test

Embodiment 1

[0023] Fabrication of a sample pretreatment device integrating protein collection and rapid enzymatic hydrolysis

[0024] 1) Fabrication of the reaction device: Drill a hole on the cap of the centrifuge tube (C), the diameter of which is equivalent to that of the solid tube a.

[0025] 2) Preparation of matrix: using glycidyl methacrylate as a monomer and ethylidene dimethacrylate as a crosslinking agent to prepare a porous monolithic matrix. Weigh 0.6g of glycidyl methacrylate (GMA), 0.4g of ethylidene dimethacrylate (EDMA), dissolve in 1g of cyclohexanol / dodecanol (mass ratio: 50 / 50) mixed solvent, ultrasonically After 10 minutes, wrap the centrifuge tube containing the reaction solution with aluminum foil. Quickly weigh 1mg of photoinitiator DMPA and shake for 30s. Such as figure 1 As shown, take 0.8mL of the reaction solution c into the reactor b, insert the solid tube a, and put it into the ultraviolet crosslinking apparatus for photopolymerization for 10 minutes.

[...

Embodiment 2

[0031] 1) Fabrication of the reaction device: Drill a hole on the cap of the centrifuge tube (C), the diameter of which is equivalent to that of the solid tube a.

[0032] 2) Preparation of matrix: using glycidyl methacrylate as a monomer and ethylidene dimethacrylate as a crosslinking agent to prepare a porous monolithic matrix. Weigh 0.6g of glycidyl methacrylate (GMA), 0.4g of ethylidene dimethacrylate (EDMA), dissolve in 1g of cyclohexanol / dodecanol (mass ratio: 50 / 50) mixed solvent, ultrasonically After 10 minutes, wrap the centrifuge tube containing the reaction solution with aluminum foil. Quickly weigh 1mg of photoinitiator DMPA and shake for 30s. Such as figure 1 As shown, take 0.8mL of the reaction solution c into the reactor b, insert the solid tube a, and put it into the ultraviolet crosslinking apparatus for photopolymerization for 10 minutes.

[0033] 3) After taking out the solid tube a, add methanol and soak in the reaction device for 4 hours, then add 29% a...

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Abstract

The invention relates to a sample pretreatment device for improving protein analysis speed, which combines protein collection and enzymatic hydrolysis functions. The invention relates to a preparation method thereof. The sample pretreatment device comprises a centrifuge tube with an upper cover, which is characterized in that the middle part of the upper cover of the centrifugal tube is provided with a hole, the inner wall of the centrifugal tube is attached with an annular hollow porous rigid integral substrate layer whose surface is fixed with enzymes via active groups modifying the substrate. The sample pretreatment device has the advantages of large sample processing amount, fast protein enzymatic hydrolysis speed, simple manufacture method and low cost, which can be used for fast enzymatic hydrolysis of collected protein samples. The invention has high practical value for the research of high flux protein.

Description

technical field [0001] The invention relates to a sample pretreatment device to improve the speed of protein analysis, that is, a sample pretreatment device integrating protein collection and enzymatic hydrolysis and its preparation. In the invention, the whole material of immobilized enzyme is synthesized in the centrifuge tube, so as to realize rapid enzymolysis at the same time of protein collection. At the same time, the device can also be used as a sample bottle for storing enzymatic hydrolysis products. Background technique [0002] During the analysis of complex protein samples, steps such as protein mixture pre-fractionation, fraction collection, free solution enzymatic digestion, peptide separation and protein identification are usually included. Fraction collection and enzymatic hydrolysis belong to the sample pretreatment process. However, free solution enzymatic hydrolysis has many disadvantages, such as long enzymatic hydrolysis time (needing more than ten hou...

Claims

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Application Information

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IPC IPC(8): G01N1/28
Inventor 张丽华袁辉明吴帅宾梁振张玉奎
Owner DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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