喉癌Hep-2细胞来源的exosomes提取方法的比较

梁俊毅, 吉晓滨, 刘启才, 等. 喉癌Hep-2细胞来源的exosomes提取方法的比较[J]. 临床耳鼻咽喉头颈外科杂志, 2015, 29(17): 1522-1526. doi: 10.13201/j.issn.1001-1781.2015.17.008
引用本文: 梁俊毅, 吉晓滨, 刘启才, 等. 喉癌Hep-2细胞来源的exosomes提取方法的比较[J]. 临床耳鼻咽喉头颈外科杂志, 2015, 29(17): 1522-1526. doi: 10.13201/j.issn.1001-1781.2015.17.008
LIANG Junyi, JI Xiaobin, LIU Qicai, et al. Comparison of methods for isolating exosomes derived from laryngocarcinoma Hep-2 cells[J]. J Clin Otorhinolaryngol Head Neck Surg, 2015, 29(17): 1522-1526. doi: 10.13201/j.issn.1001-1781.2015.17.008
Citation: LIANG Junyi, JI Xiaobin, LIU Qicai, et al. Comparison of methods for isolating exosomes derived from laryngocarcinoma Hep-2 cells[J]. J Clin Otorhinolaryngol Head Neck Surg, 2015, 29(17): 1522-1526. doi: 10.13201/j.issn.1001-1781.2015.17.008

喉癌Hep-2细胞来源的exosomes提取方法的比较

  • 基金项目:

    广东省科技厅产业技术研究与开发资金计划项目(No:2012B031800340)

    广州市科技和信息化局社会发展应用基础研究专项(No:2013J4100024)

详细信息
    通讯作者: 吉晓滨,E-mail:chusebaba@163.com
  • 中图分类号: R739.6

Comparison of methods for isolating exosomes derived from laryngocarcinoma Hep-2 cells

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  • 目的:对分离提取喉癌Hep-2细胞来源的exosomes 2种方法进行对比,展现不同方法的优缺点,为选择分离提取包括喉癌Hep-2细胞在内的肿瘤细胞来源exosomes的方法提供参考。方法:大量培养喉癌Hep-2细胞,42℃热休克处理。收集90 ml培养上清液,先通过3/0.8 μm深层过滤小型滤芯对上清液进行预处理,去除直径较大的颗粒和杂质,再利用蔗糖密度梯度离心联合超滤离心法,将上清液浓缩提纯;收集培养上清液4 ml,依次加入Exosome Isolation Kit内试剂,通过exosomes提取专用过滤柱,收集浓缩液。用高倍透射电子显微镜对2种方法所得的exosomes浓缩液分别鉴定,作出评价。结果:2种方法均能成功地从喉癌Hep-2细胞培养上清液中分离提取出exosomes。单个高倍视野下,蔗糖密度梯度离心联合超滤离心法提取exosomes分散性较好,但密度较低,背景见杂质较多;Exosome Isolation Kit所提取exosomes排列紧密,密度较高,背景杂质较少。结论:2种方法各具特点,均是较理想的exosomes分离提取方法。利用Exosome Isolation Kit分离提取exosomes具有样本量少、提取时间短、步骤简单、产物量大等优点,为喉癌Hep-2来源exosomes的分离提取提供了较好的选择。
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  • [1]

    JOHNSTONE R M,ADAM M,HAMMOND J R,et al.Vesicle formation during reticulocyte maturation.Association of plasma membrane activitive with released vesicles (exosomes)[J].J Biol Chem,1987,262:9412-9420.

    [2]

    KALANI A, TYAGI A, TYAGI N.Exosomes: mediators of neurodegeneration, neuroprotection and therapeutics[J].Mol Neurobiol,2014,49:590-600.

    [3]

    LANE R E,KORBIE D,ANDERSON W,et al.Analysis of exosome purification methods using a model liposome system and tunable-resistive pulse sensing[J].Sci Rep,2015,6:7639-7646.

    [4]

    FRANZEN C A,SIMMS P E,VAN HUIS A F,et al.Characterization of uptake and internalization of exosomes by bladder cancer cells[J].Biomed Res Int,2014,2014:619-629.

    [5]

    TAURO B J,GREENING D W,MATHIAS R A,et al.Comparison of ultracentrifugation,density gradient separation,and immunoaffinity capture methods for isolating human colon cancer cell line LIM1863-derived exosomes[J].Methods,2012,56:293-304.

    [6]

    WEBBER J,STONE T C, KATILIUS E,et al.Proteomics analysis of cancer exosomes using a novel modified aptamer-based array (SOMAscanTM) Platform[J].Mol Cell Proteomics,2014,13:1050-1064.

    [7]

    JOHNSTONE R M,MATHEW A,MASON A B,et al.Exosome formation during maturation of mammalian and avian reticulocytes:evidence that exosome release is a major route for externalization of obsolete membrane proteins[J].J Cell Physiol,1991,147:27-36.

    [8]

    ZHANG H G,GRIZZLE W E.Exosomes:a novel pathway of local and distant intercellular communication that facilitates the growth and metastasis of neoplastic lesions[J].Am J Pathol,2014,184:28-41.

    [9]

    余思,邓建中,李志澄,等.未成熟树突状细胞-胃癌细胞融合细胞来源的exosome诱导特异性抗肿瘤免疫[J].中华临床医师杂志(电子版),2013,7(2):113-116.

    [10]

    张家模,吴小候, 张翾,等.白细胞介素2锚定的exosomes的制备及其对膀胱癌细胞的诱导杀伤效应[J].中华肿瘤杂志,2011,33(8):564-569.

    [11]

    WANG J, ZHOU Y, LU J, et al. Combined detection of serum exosomal miR-21 and HOTAIR as diagnostic and prognostic biomarkers for laryngeal squamous cell carcinoma[J].Med Oncol,2014,31:148-150.

    [12]

    ZHANG Z Y, WANG C X, LI T,et al.Comparison of ultracentrifugation and density gradient separation methods for isolating Tca8113 human tongue cancer cell line-derived exosomes[J].Oncol Lett,2014,8:1701-1706.

    [13]

    TAYLOR D D,ZACHARIAS W,GERCEL-TAYLOR C.Exosomes isolation for proteomic analyses and RNA profiling[J].Methods Mol Biol,2011,728:235-246.

    [14]

    董超,刘迪,冯业童,等.结直肠癌细胞外泌体的制备及其促单核细胞增殖作用[J].吉林大学学报(医学版),2013,39(3):472-476.

    [15]

    HOOD J L,SCOTT M J,WICKLINE S A.Maximizing exosome colloidal stability following electroporation[J].Anal Biochem,2014,448:41-49.

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收稿日期:  2015-06-07

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