Isolation, Purification and Characterization of Exosomes from Fibroblast Cultures of Skeletal Muscle

被引:6
作者
van de Vlekkert, Diantha [1 ]
Qiu, Xiaohui [1 ]
Annunziata, Ida [1 ]
D'Azzo, Alessandra [1 ]
机构
[1] St Jude Childrens Res Hosp, Dept Genet, Memphis, TN 38105 USA
关键词
Fibroblasts; Cell culture; Exosomes; Ultracentrifugation; Sucrose gradient; Western;
D O I
10.21769/BioProtoc.3576
中图分类号
Q [生物科学];
学科分类号
07 ; 0710 ; 09 ;
摘要
Exosomes are dynamic nanovesicles secreted by virtually all cells and are present in all biological fluids. Given their highly heterogeneous content exosomes have been implicated in many physiological and pathological processes that they exert by influencing cell-cell and cell-ECM communication. In recent years an increasing number of methods have been established for the purification and characterization of exosomes. These include ultracentrifugation, ultrafiltration, size exclusion chromatography, immune capture and precipitation using a proprietary polymer. Here, we provide a protocol based on differential ultracentrifugation and sucrose density gradients tailored for the isolation of crude and ultra-pure exosomes from primary fibroblast cultures derived from adult mouse skeletal muscle. This protocol can be adapted and modified for the isolation and characterization of exosomes from a variety of tissues and bodily fluids.
引用
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页数:19
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