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Quantitative analysis method capable of simultaneously realizing enrichment and similar-isobar labeling of polysaccharides

A near-isobaric labeling and quantitative analysis technology, applied in the fields of systems biology and glycomics, can solve the problems of complex polysaccharide structure, no enrichment and quantitative labeling, low abundance of glycosylated proteins, etc., to achieve enhanced enrichment efficiency, improved hydrophobicity and electrospray ionization efficiency, and the effect of high-accuracy qualitative and quantitative analysis

Inactive Publication Date: 2016-11-09
TONGJI UNIV
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Problems solved by technology

Due to the very low abundance of glycosylated proteins (2%-5%) and the extremely complex structure of polysaccharides, qualitative and quantitative analysis of polysaccharides is still a great challenge
At present, there is no near-isobaric labeling quantification; at the same time, there is no report integrating enrichment and quantitative labeling

Method used

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  • Quantitative analysis method capable of simultaneously realizing enrichment and similar-isobar labeling of polysaccharides
  • Quantitative analysis method capable of simultaneously realizing enrichment and similar-isobar labeling of polysaccharides

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Embodiment

[0029] A quantitative analysis method for simultaneous enrichment and near-isobaric labeling of polysaccharides, such as figure 1 shown, including the following steps:

[0030] (1) Two groups of polysaccharides with different physiological or pathological conditions were used as the control group and the disease group respectively, and all aminations of N-acetylglucosamine aldehyde groups were labeled with the same gravity. One group of polysaccharides was labeled with sodium cyanoborohydride (NaCNBH 3 ) and H 2 15 NCH 2 COOR f labeling, another group of polysaccharides with sodium cyanoborohydride and H 2 NCH 2 13 COOR f mark, where R f for C 3 f 7 CH 2 CH 2 PhCH 2 -, respectively marked as -CH 2 15 NHCH 2 COOR f and -CH 2 NHCH 2 13 COOR f , where R f for C 3 f 7 CH 2 CH 2 PhCH 2 -, the mass difference of the polysaccharide containing the fluorine label after its labeling is 6.32mDa; Carry out the reaction process and the condition of isobaric label...

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Abstract

The invention relates to a quantitative analysis method capable of simultaneously realizing enrichment and similar-isobar labeling of polysaccharides. The method comprises the following steps: respectively using two groups of polysaccharides with different physiological or pathological conditions as a control group and a disease group, separately carrying out full amination similar-isobar labeling of a N-acetylglucosamine aldehyde group and respectively labeling the two groups of polysaccharides as -CH2<15>NHCH2COORf and -CH2<13>NHCH2COORf, wherein Rf is C3F7CH2PhCH2-, and mass difference of the labeled polysaccharides containing fluorine labels is 6.32 mDa; mixing the two groups of polysaccharides after isobar labeling in proportion and carrying out mass spectrometry so as to obtain a data set; and subjecting the data set to qualitative and quantification database search so as to obtain the topological structure of the polysaccharides and the proportion of each polysaccharide in the disease group relative to the control group, i.e., the up-regulating or down-regulating condition of all the polysaccharides under a disease condition. A quantitative labeling agent used in the method has the advantages of high labeling efficiency, high accuracy, low price and easy availability; moreover, the quantitative labeling agent has a fluorine enrichment label, so the quantitative labeling agent substantially improves the enrichment efficiency, electrospray ionization efficiency and mass spectrometric sensitivity and efficiency of the labeled polysaccharides and is applicable to quantification of N-polysaccharides based on high-resolution tandem mass spectrometric analysis.

Description

technical field [0001] The invention relates to a quantitative analysis method for simultaneous enrichment and near-isobaric labeling of polysaccharides, and mainly relates to the technical fields of systems biology and glycomics related to biological mass spectrometry. Background technique [0002] Glycosylation is one of the most extensive and important post-translational modifications of proteins; more than 50% of proteins in mammalian cells have glycosylation modifications; these modifications are closely related to many pathological processes. Due to the very low abundance of glycosylated proteins (2%-5%) and the extremely complex structure of polysaccharides, the qualitative and quantitative analysis of polysaccharides is still a huge challenge. N-glycans are the most common of the glycosylated polysaccharides. N-glycans have a core structure consisting of 2 N-acetylglucosamines and 3 mannoses; one of the N-acetylglucosamines is attached to the protein. N-glycans on ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/02G01N30/06G01N30/08G01N30/72
CPCG01N30/02G01N30/06G01N30/08G01N30/72G01N2030/027
Inventor 方后琴朱仲良田志新
Owner TONGJI UNIV
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