尿液外泌体作为重要的疾病标志物，可以为泌尿系统肿瘤的诊断和预后提供关键的生物信息。然而，目前针对尿液外泌体的分析方法主要局限于整体平均法，检测的稳定性易受外泌体总浓度波动影响。为精准分析尿液外泌体携带的分子信息，本项目拟结合团队在微流控芯片、荧光原位成像和单外泌体分析等领域的工作基础，发展基于序列化探针识别和核酸链交换信号扩增（Sequential-SABER）的单外泌体多参数荧光原位成像策略，用于尿液外泌体的分析研究。相比整体平均法，单外泌体异质性分析可以挖掘群体信息中少量肿瘤外泌体的个体异质性特征，减少由于外泌体总浓度波动造成的分析检测数据不稳定。本项目计划通过分析超过100例临床膀胱癌患者尿液样品中单外泌体水平的十余种关键蛋白表达，利用t-SNE聚类分析揭示尿液外泌体之间的个体差异，探索尿液外泌体异质性特征与临床膀胱癌的相关性，为单外泌体分析用于临床检测的研究提供理论基础和实验依据。

As an important biomarker, urine exosomes could provide key biological information for the diagnosis and prognosis of urinary tumors. However, the current methods for urine exosomes analysis are mainly limited to bulk test. Because of the fluctuations of overall exosomes concentration in urine samples, the bulk test strategy usually lacks stability for clinical applications. In order to accurately analyze the biological information carried by urine exosomes, this project plans to combine the team's research expertise in the fields of microfluidic chips, fluorescence in situ imaging and single exosomes analysis to develop a sequential-SABER based strategy for analysis of urine exosome heterogeneity. Comparing with the overall analysis method, single exosomal heterogeneity analysis can investigate the characteristics of the small portion of tumor exosomes in the whole exosome population, which can improve the detection stability. In this project, we plan to analyze the expression of more than 10 key proteins on individual urine exosomes in clinical urine samples and applied for testing of more than 100 bladder cancer patients. Besides, the t-SNE analysis method would be used to reveal the differences between individual urine exosomes in those clinical samples. We think this project would provide a theoretical and experimental basis for the applications of single exosomes heterogeneity analysis in clinical use.