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Method for in-situ desalination and enrichment on trace amount of protein or polypeptide target

A protein and trace technology, applied in the field of biochemical analysis, can solve the problems of insufficient elution, affecting detection sensitivity, and not suitable for large-scale applications of proteomics, achieving high sensitivity, high reproducibility, and improved sensitivity Effect

Inactive Publication Date: 2009-02-18
FUDAN UNIV
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Problems solved by technology

In this way, not only the water washing process requires certain experience and technology, otherwise the salt elution will be insufficient or the sample will be eluted due to excessive elution, which will not only affect the detection sensitivity but also affect the high throughput and high speed of the analysis process. Not well suited for proteomics-scale applications

Method used

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  • Method for in-situ desalination and enrichment on trace amount of protein or polypeptide target
  • Method for in-situ desalination and enrichment on trace amount of protein or polypeptide target
  • Method for in-situ desalination and enrichment on trace amount of protein or polypeptide target

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Embodiment 1

[0028] After the protein from rat liver was separated by 2D-gel and stained with Coomassie Brilliant Blue, 10 points were selected, and the labels of the points were represented by 1 to 10 according to the staining depth from deep to light ( Figure 5 ), followed by enzymolysis with trypsin. First, the disc on the MALDI target plate is coated with a block polymer of microscopic phase separation structure. After the polymer dot film is formed, 0.5 microliters of enzymatic peptide sample solution (solvent is 50% acetonitrile, 50% Water, 0.1% trifluoroacetic acid) were spotted on the coating dot, another 0.5 microliter of the same enzymatic peptide sample solution was dotted on the light target dot, and dried naturally at room temperature; finally, the volume of 0.5 microliter The organic matrix solution (2-cyano-4-hydroxycinnamic acid (α-CHCA), the solvent is 50% acetonitrile, 50% water, 0.1% trifluoroacetic acid) was spotted on the surface of the sample, and dried naturally at ...

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Abstract

The invention belongs to the biochemical analysis field and relates to a method which makes use of the block polymer coating with a micro-phase separation structure to carry out on-target elution-free in-situ desalting and synchronous enrichment on the peptide and protein samples to avoid samples loss and can directly carry out matrix-assisted laser analysis ionization mass spectrometry analysis and identification on the enriched samples. The invention can realize desalting dispensing with extra eluted desalting step and excessive matrix solution and can directly make MALDI MS analysis; the invention successfully achieves the characteristics of sample on-target in-situ elution-free desalting and enrichment at the first time and has the advantages of high sensitivity, high reproducibility, high flux and strong salt-tolerant ability, which is economical and time-saving; the invention can be widely used in the field of proteomics and also greatly expands the application scope of the polymer coating technology.

Description

technical field [0001] The invention belongs to the field of biochemical analysis and relates to a method for desalting and enriching trace samples. Specifically, it involves the use of a block polymer coating with a microscopic phase separation structure to perform in-situ desalting and simultaneous enrichment on the target without elution for polypeptide and protein samples, which can avoid sample loss and directly enrich the enriched samples. A method for performing analysis and identification by matrix-assisted laser desorption ionization mass spectrometry. Background technique [0002] With the completion of the complete sequencing of the human genome, the focus of life science research will shift from genomics to proteomics in the post-genome era. Proteome research has become one of the focuses of life science in the 21st century. The human proteome has become one of the largest strategic resources in the new century. It is the strategic commanding height and the foc...

Claims

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Application Information

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IPC IPC(8): G01N1/34G01N27/64
Inventor 张莹陆豪杰杨芃原
Owner FUDAN UNIV
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