采用金基标签的电感耦合等离子体质谱法定量小鼠血清来源的细胞外囊泡中的CD81

IF 5.6 1区化学 Q1 CHEMISTRY, ANALYTICAL

Talanta Pub Date : 2025-07-21 DOI:10.1016/j.talanta.2025.128609

Jaime Martínez-García, Beatriz Fernández, Rosario Pereiro

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引用次数: 0

摘要

细胞外囊泡（Extracellular vesicles, EVs）是细胞间通讯的主要机制之一。如今，人们知道它们在癌症或神经退行性疾病中起着重要作用。因此，开发合适的程序来分析电动汽车，以利用它们的潜力来理解生物失调和识别生物标志物，如蛋白质，是非常有意义的。然而，ev中特定蛋白质的测定通常需要高度敏感的方法，特别是在样品体积有限的情况下，如小鼠血清。金属标记免疫探针与电感耦合等离子体质谱（ICP-MS）检测相结合已被证明是分析生物样品中蛋白质的一种有利策略。在这项工作中，我们开发了一种基于免疫分析法的方法，该方法使用带有金纳米簇标记的抗体来实现高信号放大，并采用ICP-MS检测来确定小鼠血清中ev中的靶蛋白。为了验证这一概念，研究了电动汽车的通用标记CD81，获得了低至56 fM的检测限。采用优化的差速离心方案，从0.5 mL商业小鼠血清中分离出ev，测定其CD81水平。用所开发的分析方法获得的结果与商用ELISA试剂盒的结果一致。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

Quantification of CD81 in mouse serum-derived extracellular vesicles via inductively coupled plasma – mass spectrometry using gold-based labels

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Quantification of CD81 in mouse serum-derived extracellular vesicles via inductively coupled plasma – mass spectrometry using gold-based labels

Extracellular vesicles (EVs) released by cells are one of the main mechanisms of intercellular communication. Nowadays it is known that they can play important roles in diseases such as cancer or neurodegenerative processes. Therefore, it is of high interest to develop proper procedures for analyzing EVs in order to exploit their potential to understand biological dysregulations and identifying biomarkers, such as proteins. However, the determination of specific proteins in EVs typically require highly sensitive methods, particularly when working with limited sample volumes, such as mouse serum. The combined use of metal labelled immunoprobes and detection by inductively coupled plasma – mass spectrometry (ICP-MS) has proved to be an advantageous strategy for the analysis of proteins in biological samples. In this work, a methodology based on an immunoassay using antibodies labelled with gold nanoclusters, to achieve a high signal amplification, and ICP-MS detection was developed to determine target proteins in EVs from mouse blood serum. As a proof of concept, CD81, a generic marker of EVs, was investigated, and a limit of detection as low as 56 fM was obtained. CD81 levels in EVs isolated from aliquots of 0.5 mL of commercial mouse blood serum were determined after applying an optimized differential centrifugation protocol. Results obtained with the developed analytical procedure were in good agreement with those achieved with a commercial ELISA kit.

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来源期刊

Talanta 化学-分析化学

CiteScore

12.30

自引率

4.90%

发文量

861

审稿时长

29 days

期刊介绍： Talanta provides a forum for the publication of original research papers, short communications, and critical reviews in all branches of pure and applied analytical chemistry. Papers are evaluated based on established guidelines, including the fundamental nature of the study, scientific novelty, substantial improvement or advantage over existing technology or methods, and demonstrated analytical applicability. Original research papers on fundamental studies, and on novel sensor and instrumentation developments, are encouraged. Novel or improved applications in areas such as clinical and biological chemistry, environmental analysis, geochemistry, materials science and engineering, and analytical platforms for omics development are welcome. Analytical performance of methods should be determined, including interference and matrix effects, and methods should be validated by comparison with a standard method, or analysis of a certified reference material. Simple spiking recoveries may not be sufficient. The developed method should especially comprise information on selectivity, sensitivity, detection limits, accuracy, and reliability. However, applying official validation or robustness studies to a routine method or technique does not necessarily constitute novelty. Proper statistical treatment of the data should be provided. Relevant literature should be cited, including related publications by the authors, and authors should discuss how their proposed methodology compares with previously reported methods.