高等学校化学学报

高等学校化学学报

• 研究论文 • 上一篇    下一篇

外泌体处理温度对其蛋白质组学分析的影响研究

吴梦鸽1,2，楚占营2，孟  波2，叶子弘1，翟  睿2   

  1. 1. 中国计量大学生命科学学院 2.中国计量科学研究院，前沿计量科学中心

  • 收稿日期:2025-01-20 修回日期:2025-02-24 网络首发:2025-02-28 发布日期:2025-02-28
  • 通讯作者: 翟睿 E-mail:zhairui@nim.ac.cn
  • 基金资助:
    中国计量科学研究院院基本科研业务费(批准号：AKYCX2211)和科技创新领军人才计划项目(批准号：WR2202)资助

Effect of exosome treatment temperature on proteomic analysis of exosomes

WU Mengge1,2，CHU Zhanying2，MENG Bo2，YE Zihong1，ZHAI Rui2   

  1. 1. Faculty of Life Sciences, China Jiliang University 2. Center for Cutting-Edge Metrology, National Institute of Metrology,

  • Received:2025-01-20 Revised:2025-02-24 Online First:2025-02-28 Published:2025-02-28
  • Supported by:
    Supported by the Fundamental Research Funds of the National Institute of Metrology(No.AKYCX2211) and Plan for Leading Talents of Science and Technology Innovation (No.WR2202)

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摘要： 外泌体蛋白质组学分析在重大疾病生物标志物筛选、药物靶点发现及功能机制研究中具有重要价值。外泌体和蛋白质都被公认为是极易受处理温度变化影响的研究对象，然而此前的报道仍通常将基于不同富集策略得到的外泌体蛋白质组学分析结果的差异，直接归因于不同富集方法的富集选择性和富集效率的不同，关于外泌体的富集处理过程中的温度变化对外泌体蛋白质组学分析结果的影响尚未有充分的研究讨论。为探究样本处理温度对外泌体蛋白质组学分析结果的影响，本研究在4℃、25℃、37℃、45℃、60℃和90℃六种温度条件下分别处理外泌体样本1小时，随后系统分析了样本的外泌体形貌特征、颗粒粒径分布与浓度、特征蛋白质表征含量变化、以及蛋白质组学数据。结果显示，不同温度处理的外泌体在形貌和粒径上未表现出显著差异；但随着温度升高，外泌体颗粒的浓度显著下降。温度超过45℃时，外泌体标志性蛋白TSG101的表征含量显著下降，而与之相反的是，外泌体特征膜蛋白CD9的表征含量则显著上升。定量蛋白质组学分析结果进一步表明，在4℃、25℃和37℃下处理的外泌体样本的蛋白质组学数据具有较好的可比性，而45℃及以上温度处理的样本则表现出48种蛋白的显著差异。综上，在进行外泌体蛋白质组学分析时，应充分考虑温度变化对测量结果的潜在影响，以确保数据的可靠性、重现性与可比性。

关键词: 外泌体蛋白质组, 温度, 质谱

Abstract: Exosome proteomic analysis has important value in screening biomarkers for major diseases, discovering drug targets, and studying functional mechanisms. Exosome and protein are widely recognized as research objects that are highly susceptible to temperature changes. However, previous reports have often attributed the differences in exosome proteomic analysis results based on different enrichment strategies directly to the differences in enrichment selectivity and efficiency of different enrichment methods. There has not been sufficient research and discussion on the impact of temperature changes during the enrichment process of exosome on the results of exosome proteomic analysis. To investigate the effect of sample processing temperature on the results of exosome proteomics analysis, this study treated exosome samples at six temperature conditions of 4℃, 25℃, 37℃, 45℃, 60℃, and 90℃ for 1 hour. Subsequently, the morphology characteristics, particle size distribution and concentration, changes in characteristic protein characterization content, and proteomic data of the samples were systematically analyzed. The results showed that there were no significant differences in morphology and particle size among exosomes treated at different temperatures, But as the temperature increases, the concentration of exosome particles significantly decreases. When the temperature exceeds 45℃, the characterization content of the exosome marker protein TSG101 significantly decreases, while conversely, the characterization content of the exosome characteristic membrane protein CD9 significantly increases. The results of quantitative proteomics analysis further indicate that the proteomic data of exosome samples treated at 4℃, 25℃, and 37℃ have good comparability, while samples treated at 45℃ and above show significant differences in 48 proteins. In summary, when conducting exosomal proteomic analysis, the potential impact of temperature changes on measurement results should be fully considered to ensure the reliability, reproducibility, and comparability of the data.

Key words: Exosome proteomic, Temperature, Mass spectrometry

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