Mass spectrometry method for complete glycosylated peptide fragment based on quasi-multistage spectrum

A mass spectrometry detection and glycosylation technology, applied in the field of glycosylation proteomics, can solve the problems of ion difficulty, glycopeptide analysis throughput, limited sensitivity and identification efficiency, and achieve the effect of improving specificity

Active Publication Date: 2019-07-02
DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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Problems solved by technology

This method achieves the simultaneous fragmentation of sugar chains and peptide backbones, but it is difficult to simultaneously screen ions for further fragmentation due to the impact of ion transport efficiency
The abov

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  • Mass spectrometry method for complete glycosylated peptide fragment based on quasi-multistage spectrum
  • Mass spectrometry method for complete glycosylated peptide fragment based on quasi-multistage spectrum
  • Mass spectrometry method for complete glycosylated peptide fragment based on quasi-multistage spectrum

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Embodiment 1

[0025] The mouse tissues / human liver cancer tissues and paracancerous tissues used in this experiment were provided by the Second Affiliated Hospital of Dalian Medical University (Dalian, China). The acquisition and use of the samples were completely legal and complied with the relevant regulations of the ethics committee of the hospital. The experimental procedure is as follows:

[0026] 1. Preparation of protein samples: take 50-100 mg of human tissue, clean it with normal saline on ice, cut it into pieces, and wash it with ice normal saline, then grind it into powder under liquid nitrogen, add 5-10 mL Protein lysate, protein is extracted by ultrasonic crushing method. The protein lysate consists of: 6M guanidine hydrochloride, 50mM Tris‐HCl (pH 8.0), 20mM DTT. Centrifuge the protein lysate at high speed, take the supernatant and add it to 5 times the volume of the protein pre-cooled precipitation solution (acetone: ethanol: acetic acid = 50:50:0.1, v / v / v), and place it in...

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Abstract

The invention relates to a mass spectrometry method for a complete glycosylated peptide fragment based on quasi-multistage spectrum and an application thereof in early diagnosis of liver cancer. The method mainly comprises the following five steps: 1) enrichment of a glycosylated peptide fragment; 2) sample preparation of a deglycosylated peptide fragment-complete glycosylated peptide fragment; 3)mass spectrometry of the deglycosylated peptide fragment-complete glycosylated peptide fragment; 4) spectrogram data retrieval of the deglycosylated peptide fragment-complete glycosylated peptide fragment; and 5) based on the above, through combination of glycosylation site and complete glycopeptide quantification technology, realizing glycosylation refined difference analysis of a cancer patientand a normal person and screening potential disease markers. The method realizes refined analysis of human protein glycosylation, and has important application potential in screening the tumor markers.

Description

technical field [0001] The invention belongs to the technical field of glycosylation proteomics in the research direction of proteomics, and specifically relates to the high-throughput identification and quantitative analysis of N-linked glycosylated peptides, and the application of the technology in clinical diagnosis of tumors. Background technique [0002] As the most common and important post-translational modification, protein glycosylation is involved in immune response, cell-cell interaction, ligand-receptor interaction and other processes, and its abnormality is closely related to the occurrence of diseases. At present, most of the protein tumor markers used clinically are glycosylated proteins, such as the marker protein of liver cancer - alpha-fetoprotein (AFP), the marker of malignant tumor - cancer antigen 125 (cancer antigen 125), and the marker of prostate cancer Substances-prostate cancer specific antigen (PSA) etc. However, the current tumor markers mainly f...

Claims

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Application Information

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IPC IPC(8): G01N27/62
CPCG01N33/6848
Inventor 叶明亮秦洪强陈尧王璐
Owner DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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