Luo Heyong, Zhang Jing, Yang Anqing, Ouyang Weiwei, Long Shiqi, Lin Xiaojin, Yang Na, Yang Zhiru, Zhang Yingchun, Yang Wei, Che Qiyuan, Yang Yuxin, Guo Ting, Zhao Xing
Center for Tissue Engineering and Stem Cell Research, Guizhou Medical University, Guiyang, China.
Department of Immunology, College of Basic Medical Sciences, Guizhou Medical University, Guiyang, China.
Bio Protoc. 2023 Jun 5;13(11):e4693. doi: 10.21769/BioProtoc.4693.
Exosomes are lipid bilayer-enclosed vesicles, actively secreted by cells, containing proteins, lipids, nucleic acids, and other substances with multiple biological functions after entering target cells. Exosomes derived from NK cells have been shown to have certain anti-tumor effects and potential applications as chemotherapy drug carriers. These developments have resulted in high demand for exosomes. Although there has been large-scale industrial preparation of exosomes, they are only for generally engineered cells such as HEK 293T. The large-scale preparation of specific cellular exosomes is still a major problem in laboratory studies. Therefore, in this study, we used tangential flow filtration (TFF) to concentrate the culture supernatants isolated from NK cells and isolated NK cell-derived exosomes (NK-Exo) by ultracentrifugation. Through a series of characterization and functional verification of NK-Exo, the characterization, phenotype, and anti-tumor activity of NK-Exo were verified. Our study provides a considerably time- and labor-saving protocol for the isolation of NK-Exo.
外泌体是由脂质双分子层包裹的囊泡，由细胞主动分泌，进入靶细胞后含有具有多种生物学功能的蛋白质、脂质、核酸和其他物质。来自自然杀伤细胞（NK细胞）的外泌体已被证明具有一定的抗肿瘤作用，并作为化疗药物载体具有潜在应用价值。这些进展导致对外泌体的需求很高。尽管外泌体已有大规模工业制备，但仅针对诸如人胚肾293T等一般工程细胞。特定细胞外泌体的大规模制备仍是实验室研究中的一个主要问题。因此，在本研究中，我们使用切向流过滤（TFF）浓缩从NK细胞分离的培养上清液，并通过超速离心分离NK细胞衍生的外泌体（NK-Exo）。通过对NK-Exo的一系列表征和功能验证，验证了NK-Exo的表征、表型和抗肿瘤活性。我们的研究为NK-Exo的分离提供了一种相当节省时间和人力的方案。
