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题名: An integrated sample pretreatment platform for quantitative N-glycoproteome analysis with combination of on-line glycopeptide enrichment, deglycosylation and dimethyl labeling
作者: Weng, Yejing1, 2;  Qu, Yanyan1, 2;  Jiang, Hao1, 2;  Wu, Qi1, 2;  Zhang, Lihua1;  Yuan, Huiming1;  Zhou, Yuan1, 2;  Zhang, Xiaodan1;  Zhang, Yukui1
关键词: Relative quantification ;  N-glycoproteomes ;  Integrated platform ;  N-2-assisted interface ;  Tumor metastasis
刊名: ANALYTICA CHIMICA ACTA
发表日期: 2014-06-23
DOI: 10.1016/j.aca.2014.04.037
卷: 833, 页:1-8
收录类别: SCI
文章类型: Article
WOS标题词: Science & Technology ;  Physical Sciences
类目[WOS]: Chemistry, Analytical
研究领域[WOS]: Chemistry
英文摘要: Relative quantification of N-glycoproteomes shows great promise for the discovery of candidate biomarkers and therapeutic targets. The traditional protocol for quantitative analysis of glycoproteomes is usually off-line performed, and suffers from long sample preparation time, and the risk of sample loss or contamination due to manual manipulation. In this study, a novel integrated sample preparation platform for quantitative N-glycoproteome analysis was established, with combination of online N-glycopeptide capture by a HILIC column, sample buffer exchange by a N-2-assisted HILIC-RPLC interface, deglycosylation by a hydrophilic PNGase F immobilized enzymatic reactor (hIMER) and solid dimethyl labeling on a C18 precolumn. To evaluate the performance of such a platform, two equal aliquots of immunoglobulin G (IgG) digests were sequentially pretreated, followed by MALDI-TOF MS analysis. The signal intensity ratio of heavy/light (H/L) labeled deglycosylated peptides with the equal aliquots was 1.00 (RSD = 6.2%, n = 3), much better than those obtained by the offline protocol, with H/L ratio as 0.76 (RSD = 11.6%, n = 3). Additionally, the total on-line sample preparation time was greatly shortened to 160 min, much faster than that of offline approach (24 h). Furthermore, such an integrated pretreatment platform was successfully applied to analyze the two kinds of hepatocarcinoma ascites syngeneic cell lines with high (Hca-F) and low (Hca-P) lymph node metastasis rates. For H/L labeled Hca-P lysates with the equal aliquots, 99.6% of log2 ratios (H/L) of quantified glycopeptides ranged from -1 to 1, demonstrating high accuracy of the developed sample preparation strategy. By triplicated analysis of glycopeptides and non-glycopeptides of Hca-F and Hca-P lysates, 43 up-regulated and 30 down-regulated (Hca-F/P)N-glycosylation sites, and 11 significantly changed N-glycoproteins were successfully quantified, and most of them were related to tumorigenesis and tumor metastasis. All these results demonstrate the developed integrated N-glycoprotein pretreatment platform is of great power for the accurate, precise and high-throughput analysis of N-glycoproteomes. (C) 2014 Elsevier B.V. All rights reserved.
关键词[WOS]: HYDROPHILIC-INTERACTION CHROMATOGRAPHY ;  MASS-SPECTROMETRY ;  LIQUID-CHROMATOGRAPHY ;  CANCER-CELLS ;  PROTEOME ANALYSIS ;  CARCINOMA CELLS ;  GLYCOSYLATION ;  IDENTIFICATION ;  METASTASIS ;  SEPARATION
语种: 英语
WOS记录号: WOS:000337878800001
Citation statistics: 
内容类型: 期刊论文
URI标识: http://cas-ir.dicp.ac.cn/handle/321008/145713
Appears in Collections:中国科学院大连化学物理研究所_期刊论文

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作者单位: 1.Chinese Acad Sci, Dalian Inst Chem Phys, Natl Chromatog Res & Anal Ctr, Key Lab Separat Sci Analyt Chem, Dalian 116023, Peoples R China
2.Univ Chinese Acad Sci, Beijing 100039, Peoples R China

Recommended Citation:
Weng, Yejing,Qu, Yanyan,Jiang, Hao,et al. An integrated sample pretreatment platform for quantitative N-glycoproteome analysis with combination of on-line glycopeptide enrichment, deglycosylation and dimethyl labeling[J]. ANALYTICA CHIMICA ACTA,2014,833:1-8.
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