CD63-snorkel tagging for isolation of exosomes

Extracellular vesicle Pub Date : 2023-10-26 DOI:10.1016/j.vesic.2023.100031

Chaoshan Han , Junjie Yang , Tingting Yin , Junqing An , Aijun Qiao , Yangpo Cao , Yuliang Feng , Haocheng Lu , Ying Wang , Liang Yang , Gangjian Qin

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Abstract

Exosomes (Exo) are important mediators of inter-cellular communications; however, no effective method is available for isolating, thus characterizing, cellular-specific exosomes in vivo. Since CD63 is a reliable marker for exosomes, we have developed a tagging strategy, term “CD63-Snorkel (CD63-SNKL)”, in which CD63 at its intracellular C-terminus was fused to a fragment of PDGFRB that contains the transmembrane domain tethered to multiple epitope tags (HA, His, and FLAG) displayed in tandem on surface. We found that the CD63-SNKL protein has similar subcellular localizations as endogenous CD63 and can be effectively sorted into Exo. Furthermore, Exo secreted from CD63-SNKL–transduced cells can be effectively captured on anti-HA magnetic beads and eluted with HA peptides. Thus, CD63-SNKL may be engineered for isolating and tracking endogenous tissue-specific Exo in vivo.

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CD63-snorkel标记法分离外泌体

外泌体(Exo)是细胞间通讯的重要介质;然而，没有有效的方法可用于分离，从而在体内表征细胞特异性外泌体。由于CD63是外泌体的可靠标记，我们开发了一种标记策略，称为“CD63- snorkel (CD63- snkl)”，其中细胞内c端的CD63与PDGFRB片段融合，该片段包含与多个表位标签(HA, His和FLAG)串联在表面上的跨膜结构域。我们发现CD63- snkl蛋白与内源性CD63具有相似的亚细胞定位，并且可以有效地分类到Exo中。此外，从cd63 - snkl转导的细胞中分泌的Exo可以有效地捕获在抗HA磁珠上，并用HA肽洗脱。因此，CD63-SNKL可能被设计用于体内分离和跟踪内源性组织特异性外显子。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Extracellular vesicle Biochemistry, Genetics and Molecular Biology (General)

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审稿时长

43 days