肝细胞癌 (HCC) 是一种常见的原发性肝癌，通常与慢性乙型肝炎病毒感染 (CHB) 和肝硬化 (LC) 相关，这强调了发现生物标志物以改善患者预后的迫切需要。利用小细胞外囊泡 (sEV) 液体活检的蛋白质组学技术作为生物标志物开发的一种有前景的途径，提供了新的见解。在这里，我们评估了多种 sEV 分离方法，并确定多糖壳聚糖 (CS) 是最佳方法。随后，我们采用优化的基于 CS 的磁珠 (Mag-CS) 从健康对照、CHB、LC 和 HBV-HCC 患者的血清样本中分离 sEV。利用数据独立的采集质谱与机器学习相结合，我们发现了潜在的囊泡蛋白生物标志物特征（KNG1、F11、KLKB1、CAPNS1、CDH1、CPN2、NME2），能够区分 HBV-HCC 与 CHB、LC 和非 HCC 病症。总的来说，我们的研究结果强调了基于 Mag-CS 的 sEV 分离在识别 HBV-HCC 早期检测生物标志物方面的实用性。© 2024 作者。 《Journal of Extracellular Vesicles》由 Wiley periodicals LLC 代表国际细胞外囊泡学会出版。

Hepatocellular carcinoma (HCC) is a prevalent primary liver cancer often associated with chronic hepatitis B virus infection (CHB) and liver cirrhosis (LC), underscoring the critical need for biomarker discovery to improve patient outcomes. Emerging as a promising avenue for biomarker development, proteomic technology leveraging liquid biopsy from small extracellular vesicles (sEV) offers new insights. Here, we evaluated various methods for sEV isolation and identified polysaccharide chitosan (CS) as an optimal approach. Subsequently, we employed optimized CS-based magnetic beads (Mag-CS) for sEV separation from serum samples of healthy controls, CHB, LC, and HBV-HCC patients. Leveraging data-independent acquisition mass spectrometry coupled with machine learning, we uncovered potential vesicular protein biomarker signatures (KNG1, F11, KLKB1, CAPNS1, CDH1, CPN2, NME2) capable of distinguishing HBV-HCC from CHB, LC, and non-HCC conditions. Collectively, our findings highlight the utility of Mag-CS-based sEV isolation for identifying early detection biomarkers in HBV-HCC.© 2024 The Author(s). Journal of Extracellular Vesicles published by Wiley Periodicals LLC on behalf of International Society for Extracellular Vesicles.