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Method for enriching, desalting protein or polypeptide in minute quantities, and carrying out analysis directly

A protein and enrichment technology, applied in the field of biochemical analysis and identification, can solve problems such as limited desalting ability, achieve the effect of high concentration efficiency, strong desalting ability, ensure reproducibility and reliability of results

Inactive Publication Date: 2006-12-06
FUDAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

At present, the most commonly used solvent evaporation method will also enrich the impurities such as salt while concentrating the sample, while the traditional desalination method uses the C 18 Packing ZipTip TM Nozzle desalting or on-target elution of samples with deionized water, but both methods have limited desalting capabilities

Method used

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  • Method for enriching, desalting protein or polypeptide in minute quantities, and carrying out analysis directly
  • Method for enriching, desalting protein or polypeptide in minute quantities, and carrying out analysis directly
  • Method for enriching, desalting protein or polypeptide in minute quantities, and carrying out analysis directly

Examples

Experimental program
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Effect test

Embodiment 1

[0026] Example 1, the enrichment and MALDI TOF / MS determination of silica nanoparticles to protein solution

[0027] Take 1mL concentration 10 -8 Add 2 μL of 10 mg / mL silica nanosuspension to the ubiquitin protein sample solution of M, shake at 37°C for 30 min; suspended.

[0028] Take 0.5 μL of the aforementioned suspension and drop it on a MALDI rake plate, then add an equal volume of matrix solution, and perform mass spectrometry analysis after the crystals are dried. The mass spectrometer used is MALDI TOF / TOF (4700 Proteomics Analyzer, Applied Biosystems); the laser is Nd-YAG laser, the wavelength is 355nm, and the laser pulse frequency is 200Hz; the accelerating voltage is 20kV, and the positive ion mode linear TOF detection, the detection results are shown in image 3 .

Embodiment 2

[0029] Example 2, Silica Nanoparticles Anti-salt Enrichment and MALDITOF / MS Determination of Protein Samples Containing High Concentration Inorganic Salts

[0030] Take 1 mL of Cytochrome C protein sample solution containing 4M NaCl at a concentration of Cytochrome C of 10 -7 M, add silicon oxide nano-suspension according to the method of Example 1, sample after centrifugation, positive ion mode linear TOF detection, the detection results are shown in Figure 5 .

Embodiment 3

[0031] Example 3, Silica Nanoparticles Anti-salt Enrichment and MALDITOF / MS Determination of Protein Samples Containing High Concentration Inorganic Salts

[0032] Take 1 mL of Cytochrome C protein sample solution containing 8M urea at a concentration of Cytochrome C of 10 -7 M, add silicon oxide nano-suspension according to the method of Example 1, sample after centrifugation, positive ion mode linear TOF detection, the detection results are shown in Figure 7 .

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Abstract

The invention relates to a method to directly take substrate assistant laser MALDI / MS analysis to trace protein and the protein enzymolysis sample desalination enrichment that adopts silicon oxide nanometer particle as adsorbent. It conquers the disturbance from high thickness salt and has good adsorbing effect to protein and protein enzymolysis polypeptide. The method has good desalination function. The silicon oxide has good compatibility with MALDI-MS, so it could take directly analysis and easy to operate, avoid the sample consumption. It could take identification to the protein enzymolysis sample of the thickness 10-10M, and conquers the disturbance from 4M sodium chloride or 8M urea to mass spectrum. The enrichment efficiency could reach 100 times.

Description

technical field [0001] The invention belongs to the technical field of biochemical analysis and identification, and in particular relates to a method for desalting, enriching and identifying and analyzing trace proteins or polypeptides. Utilizing the high-efficiency anti-salt effect of silica nanoparticles and its ability to enrich protein / peptide molecules, it can realize efficient desalting and enrichment of trace proteins / peptides, as well as direct matrix-assisted laser desorption ionization / mass spectrometry analysis and identification. Background technique [0002] Matrix-assisted laser desorption ionization / mass spectrometry (MALDI / MS) has become an effective tool in proteomics research due to its fast analysis speed, high sensitivity, and easy operation. However, a large amount of salt or other non-protein components (added during sample pre-treatment) in the analyzed sample will greatly reduce the sensitivity of mass spectrometry, which is called the suppression eff...

Claims

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Application Information

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IPC IPC(8): G01N30/02G01N30/08G01N1/34
Inventor 于锡娟张亚红王晓燕唐颐杨芃原
Owner FUDAN UNIV
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