Method for eluting polypeptide adsorbed to surface of meso-porous silicon material

A technology of surface adsorption and mesoporous silicon, applied in the field of peptide elution, can solve the problems of poor elution effect and low recovery rate of peptides, and achieve the effect of improving elution effect, increasing recovery rate and good compatibility

Active Publication Date: 2015-06-10
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

[0004] In order to solve the problem of poor elution effect of polypeptides in the prior art, the present invention provides a method for eluting polypeptides adsorbed on the surface of mesoporous silicon materials. Peptides are eluted
The method is simple and reliable, and effectively solves the problem of low recovery rate when extracting peptides from mesoporous silicon materials

Method used

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  • Method for eluting polypeptide adsorbed to surface of meso-porous silicon material
  • Method for eluting polypeptide adsorbed to surface of meso-porous silicon material
  • Method for eluting polypeptide adsorbed to surface of meso-porous silicon material

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] Embodiment 1: MCM-41 extracts the elution method of acidic polypeptide sample

[0025] 1) Peptide standard sample preparation with pH=3: Dissolve the mixed standard sample of 8 kinds of peptides (pep1-8) in 20mM phosphate buffer solution (pH=3) to obtain a standard sample with a concentration of 1.125pmol / μL.

[0026] 2) Peptide extraction: take 500μL of 10mg / mL mesoporous silicon material MCM-41 (pore diameter: 3.5nm, specific surface area: 800m 2 / g) suspension in a centrifuge tube, after centrifugation, remove the supernatant. Add 500 μL of the above peptide standard sample, shake and centrifuge. The material was then washed twice with 500 μL of pure water.

[0027] 3) Elution of the loaded peptide: the material adsorbed by the peptide was washed with 500 μL of 50% ACN, 1M NaCl, ACN / 100mM NaCl (1:1, v / v) and ACN / 100mM NH 4 HCO 3 (1:1, v / v) (100mM NH 4 HCO 3 pH value of 8.0) for elution. The eluate was reconstituted after being blown dry with nitrogen, and anal...

Embodiment 2

[0029] Embodiment 2: MCM-41 extracts the elution method of neutral polypeptide sample

[0030] 1) Preparation of peptide standard sample with pH=7: Dissolve the mixed standard of the above 8 peptides with 20mM phosphate buffer solution (pH=7) to obtain a standard sample with a concentration of 1.125pmol / μL.

[0031] 2) Peptide extraction: take 500μL of 10mg / mL mesoporous silicon material MCM-41 (pore diameter: 3.5nm, specific surface area: 800m 2 / g) suspension in a centrifuge tube, after centrifugation, remove the supernatant. Add 500 μL of the above peptide standard sample, shake and centrifuge. The material was then washed twice with 500 μL of pure water.

[0032] 3) Elution of the loaded peptide: the material adsorbed by the peptide was washed with 500 μL of 50% ACN, 1M NaCl, ACN / 100mM NaCl (1:1, v / v) and ACN / 100mM NH 4 HCO 3 (1:1, v / v) for elution. The eluate was reconstituted after being blown dry with nitrogen, and analyzed by LC-MS / MS.

[0033] 4) Analysis results:...

Embodiment 3

[0034] Embodiment 3: SBA-15 extracts the elution method of acidic polypeptide sample

[0035] 1) Preparation of peptide standard samples with pH=3: Dissolve the mixed standard of the above 8 peptides with 20mM phosphate buffer solution (pH=3) to obtain a standard sample with a concentration of 1.125pmol / μL.

[0036] 2) Peptide extraction: Take 500μL of 10mg / mL mesoporous silicon material SBA-15 (pore diameter: 8nm, specific surface area: 650m 2 / g) suspension in a centrifuge tube, after centrifugation, remove the supernatant. Add 500 μL of the above peptide standard sample, shake and centrifuge. The material was then washed twice with 500 μL of pure water.

[0037] 3) Elution of loaded peptide: the material with adsorbed peptide was washed with ACN / 100mM NH 4 HCO 3 (1:1, v / v) for elution. The eluate was reconstituted after being blown dry with nitrogen, and analyzed by LC-MS / MS.

[0038] 4) Analysis results: After SBA-15 extracts the peptide standard sample with pH = 3, ...

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Abstract

The invention relates to a method for eluting polypeptide adsorbed to the surface of a meso-porous silicon material. The method is characterized by comprising the step of eluting polypeptide adsorbed to the surface of the meso-porous silicon material by virtue of a mixed solution of organic solvents and a saline solution, wherein the pH value of the saline solution is more than 7 and is not less than two pH units of an isoelectric point of polypeptide with strongest alkalinity. The method is simple and reliable, and the problem that the recovery rate is low when polypeptide is extracted from the meso-porous silicon material is effectively solved.

Description

technical field [0001] The invention relates to a polypeptide elution method, in particular to a method for separating the polypeptide adsorbed on the surface of the mesoporous silicon material from the mesoporous silicon material. Background technique [0002] Most of the endogenous peptides in plasma are the products of enzymatic hydrolysis reactions in the tissue or cell microenvironment of the organism, which reflect the physiological and pathological state of the body, and are a class of potential disease markers, which have great potential in the direction of disease diagnosis and clinical treatment. Significance. However, the content of most endogenous peptides in plasma is extremely low, for example, the concentration of interleukin 6 is 0.5 pg / mL (Mol. Cell Proteomics, 2002, 1,845), which is far beyond the detection capability of current mass spectrometry. In addition, high levels of large proteins (such as serum albumin at a concentration of 35-50 mg / mL) and salts...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K1/14
Inventor 杜燕吴大朋关亚风
Owner DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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