Muraoka Satoshi, Hirano Masayo, Isoyama Junko, Ishida Mimiko, Tomonaga Takeshi, Adachi Jun
Laboratory of Proteomics for Drug Discovery, Center for Drug Design Research, National Institute of Biomedical Innovation, Health and Nutrition, Osaka 567-0085, Japan.
Laboratory of Clinical and Analytical Chemistry, Collaborative Research Center for Health and Medicine, National Institute of Biomedical Innovation, Health and Nutrition, Osaka 567-0085, Japan.
ACS Omega. 2022 Nov 3;7(45):41472-41479. doi: 10.1021/acsomega.2c05244. eCollection 2022 Nov 15.
Extracellular vesicles (EVs) are ubiquitously secreted by almost every cell type and are present in all body fluids. Blood-derived EVs can be used as a promising source for biomarker monitoring in disease. EV proteomics is currently being analyzed in clinical specimens. However, their EV proteomics preparation methods are limited in throughput for human subjects. Here, we introduced a novel automated EV isolation and sample preparation method using a magnetic particle processing robot for automated 96-well processing of magnetic particles for EV proteomics analysis that can be started with a low volume of multiple clinical samples. The automation of EV purification reduced the coefficient of variation of protein quantification from 3.5 to 2.2% compared with manual purification, enabling the quantification of 1120 proteins in 1 h of MS analysis. This automated proteomics EV sample preparation is attractive for processing large cohort samples for biomarker development, validation, and routine testing.
细胞外囊泡（EVs）几乎由所有细胞类型普遍分泌，并存在于所有体液中。血液来源的细胞外囊泡可作为疾病生物标志物监测的一个有前景的来源。目前正在临床标本中分析细胞外囊泡蛋白质组学。然而，它们的细胞外囊泡蛋白质组学制备方法在用于人类受试者时通量有限。在此，我们介绍了一种新颖的自动化细胞外囊泡分离和样品制备方法，该方法使用磁珠处理机器人对磁珠进行自动化96孔处理，用于细胞外囊泡蛋白质组学分析，可从少量多种临床样本开始。与手动纯化相比，细胞外囊泡纯化的自动化将蛋白质定量的变异系数从3.5%降低到2.2%，从而能够在1小时的质谱分析中定量1120种蛋白质。这种自动化蛋白质组学细胞外囊泡样品制备对于处理大量队列样本以进行生物标志物开发、验证和常规检测具有吸引力。
