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目的:从MDA-MB-231乳腺癌细胞培养上清中分离获得Exosomes,并进行形态学观察鉴定。方法:通过低速离心、高速离心、超滤及超高速离心等方法,从MDA-MB-231乳腺癌细胞培养上清中分离Exosomes,用透射电子显微镜观察其形态特征,Lorry方法蛋白定量,通过用免疫印记进行MHC分子鉴定。结果:MDA-MB-231乳腺癌细胞上清中分泌大量的Exosomes,电镜下观察呈椭圆形或圆形的双层膜的囊泡,直径为50～100nm,具有完整的细胞膜。结论:用离心的方法可从MDA-MB-231乳腺癌细胞培养上清中提取Exosomes,有可能作为乳腺癌免疫治疗的潜在抗原。
OBJECTIVE: To isolate Exosomes from the culture supernatant of MDA-MB-231 breast cancer cells and to observe the morphological changes. METHODS: Exosomes were isolated from the culture supernatant of MDA-MB-231 breast cancer cells by low-speed centrifugation, high-speed centrifugation, ultrafiltration and ultracentrifugation. Morphological characteristics were observed by transmission electron microscopy. The Lorry protein was quantified. Western blot MHC molecular identification. Results: Exosomes were secreted in the supernatant of MDA-MB-231 breast cancer cells, and the vesicles with oval or circular bilayers were observed under electron microscope. The diameter of the vesicles was 50-100 nm with a complete cell membrane. Conclusion: Exosomes can be extracted from the culture supernatant of MDA-MB-231 breast cancer cells by centrifugation, which may be used as potential antigen for immunotherapy of breast cancer.