纳升液相色谱-高分辨串联质谱研究不同分离方法对唾液外泌体蛋白质组的影响

引用本文: 曾琼兰, 吴利, 李水明, 王勇. 纳升液相色谱-高分辨串联质谱研究不同分离方法对唾液外泌体蛋白质组的影响[J]. 分析化学, 2018, 46(9): 1408-1414. doi: 10.11895/j.issn.0253-3820.181261 shu

Citation: ZENG Qiong-Lan, WU Li, LI Shui-Ming, WANG Yong. Comparison of Separation Methods for Isolation in Analysis of Human Saliva-derived Exosomes Proteomics by Nano-liquid Chromatography-Mass Spectrometry[J]. Chinese Journal of Analytical Chemistry, 2018, 46(9): 1408-1414. doi: 10.11895/j.issn.0253-3820.181261 shu

纳升液相色谱-高分辨串联质谱研究不同分离方法对唾液外泌体蛋白质组的影响

曾琼兰 ^1, ,
吴利 ^1, ,
李水明 ^2, ,
王勇 ^2,

1.
深圳大学生命科学学院, 深圳市海洋生物资源与生态环境重点实验室, 深圳 518060;
2.
深圳大学生命科学学院, 深圳市微生物基因工程重点实验室, 深圳 518060
基金项目:
本文系深圳市科技计划项目（No.JCYJ20170818142154551）资助

摘要: 外泌体是一系列胞外囊泡，是生物标志物的来源，但目前尚无灵敏高效的唾液外泌体蛋白质分离方法。本研究采用质谱方法比较唾液外泌体试剂盒（ExoQuick，EQ）方法和超高速离心（Ultracentrifugation，UC）方法分离外泌体的效果以及尿素缓冲液、RIPA裂解液、SDS裂解液提取外泌体蛋白质的效果。Brodford法和BCA定量测定结果表明，EQ方法分离0.5 mL唾液外泌体得到的蛋白质含量高于UC方法分离2 mL唾液所得到的蛋白质。进一步的质谱分析表明，前者鉴定到的蛋白质数目亦多于后者；试剂盒分离唾液外泌体与尿素缓冲液提取外泌体蛋白质的方法联用效果最佳，鉴定到194种蛋白质。本方法重现性好、样品用量少、检测结果稳定，可为在唾液外泌体中筛选疾病的标志物提供方法学参考。

关键词:

English

Comparison of Separation Methods for Isolation in Analysis of Human Saliva-derived Exosomes Proteomics by Nano-liquid Chromatography-Mass Spectrometry

1.
College of Life and Ocean Science, Shenzhen Key Laboratory of Marine Bioresources and Ecology, Shenzhen University, Shenzhen 518060, China;
2.
College of Life and Ocean Science, Shenzhen Key Laboratory of Microbial Genetic Engineering, Shenzhen 518060, China
Received Date: 19 April 2018
Revised Date: 19 July 2018
Fund Project:
This work was supported by the Scientific Plan Program of Shenzhen, China (No. JCYJ20170818142154551)

Abstract: Exosomes are a series of extracellular vesicles, which are the source of biomarkers. However, there are no sensitive and efficient methods for the isolation of saliva exosome proteins. In this work, we explored the effect of isolation of saliva exosome by ExoQuick (EQ) and ultracentrifugation (UC) method, and the effect of UREA buffer, RIPA lysate, and SDS lysate on exosome protein extraction by nano-liquidchromatography-high-resolution tandem mass spectrometry. The quantitative results of Brodford method showed that the protein content of exosomes obtained by EQ method from 0.5 mL of saliva was higher than that by UC method from 2 mL of saliva. Further mass spectrometry analysis showed that the former identified more proteins than the latter. The effect of the combination of using EQ method and UREA buffer to obtain saliva exosomes proteins was the best. With this method, a total of 194 proteins were identified. The method had good reproducibility, low sample requirement, and stable mass spectrometry detection, and could provide methodological support for finding markers of disease in saliva exosomes.

Key words:

Nano liquid chromatography-high resolution tandem mass spectrometry
/ Saliva-exosome
/ ExoQuick
/ Ultracentrifugation

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沈阳化工大学材料科学与工程学院沈阳 110142

纳升液相色谱-高分辨串联质谱研究不同分离方法对唾液外泌体蛋白质组的影响

English

Comparison of Separation Methods for Isolation in Analysis of Human Saliva-derived Exosomes Proteomics by Nano-liquid Chromatography-Mass Spectrometry

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中国化学会秘书处

纳升液相色谱-高分辨串联质谱研究不同分离方法对唾液外泌体蛋白质组的影响

English

Comparison of Separation Methods for Isolation in Analysis of Human Saliva-derived Exosomes Proteomics by Nano-liquid Chromatography-Mass Spectrometry

CCS Chemistry：嵌段共聚物自组装成 “三明治”二维有机材料，实现纳米级压力传感功能

CCS Chemistry：颜徐州、鲍哲南： 你的皮肤可能是一片SPMs

CCS Chemistry：工作高效不疲劳，只须让催化剂动起来

CCS Chemistry：静电组装导向聚合- 聚电解质纳米凝胶量化制备新策略

CCS Chemistry：金黄色葡萄球菌醛基脱氢酶是如何识别特异性底物的？

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