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An integrated sample pretreatment platform for quantitative N-glycoproteome analysis with combination of on-line glycopeptide enrichment, deglycosylation and dimethyl labeling
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
2014-06-23
DOI10.1016/j.aca.2014.04.037
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
关键词[WOS]HYDROPHILIC-INTERACTION CHROMATOGRAPHY ; MASS-SPECTROMETRY ; LIQUID-CHROMATOGRAPHY ; CANCER-CELLS ; PROTEOME ANALYSIS ; CARCINOMA CELLS ; GLYCOSYLATION ; IDENTIFICATION ; METASTASIS ; SEPARATION
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.
语种英语
WOS记录号WOS:000337878800001
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被引频次:14[WOS]   [WOS记录]     [WOS相关记录]
文献类型期刊论文
条目标识符http://cas-ir.dicp.ac.cn/handle/321008/145713
专题中国科学院大连化学物理研究所
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
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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.
APA Weng, Yejing.,Qu, Yanyan.,Jiang, Hao.,Wu, Qi.,Zhang, Lihua.,...&Zhang, Yukui.(2014).An integrated sample pretreatment platform for quantitative N-glycoproteome analysis with combination of on-line glycopeptide enrichment, deglycosylation and dimethyl labeling.ANALYTICA CHIMICA ACTA,833,1-8.
MLA Weng, Yejing,et al."An integrated sample pretreatment platform for quantitative N-glycoproteome analysis with combination of on-line glycopeptide enrichment, deglycosylation and dimethyl labeling".ANALYTICA CHIMICA ACTA 833(2014):1-8.
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