从小鼠股骨骨髓中分离免疫细胞

Xiaoyu Liu; Ning Quan

doi:10.21769/BioProtoc.1631

Improve Research Reproducibility A Bio-protocol resource

提交稿件
订阅
登录
/
注册
- 个人主页
- 编辑个人信息
- 修改密码
- 退出
CN
- EN - English
- CN - 中文

Peer-reviewed

Immune Cell Isolation from Mouse Femur Bone Marrow

从小鼠股骨骨髓中分离免疫细胞

Xiaoyu Liu

NQ Ning Quan email

发布: 2015年10月20日第5卷第20期 DOI: 10.21769/BioProtoc.1631 浏览次数: 24552

评审: Oneil G. BhalalaThomas X. LuAnonymous reviewer(s)

PDF

Q&A

引用

Cited by

参见作者原研究论文

The authors used this protocol in:

Cover of The Journal of Neuroscience, featuring study using the protocol.

Feb 2015

实验方案合集

Cell Imaging - A Special Collection for Cell Bio 2023

Abstract

The bone marrow is the site of hematopoiesis and contains mixed population of blood cells including erythrocytes, granulocytes, monocytes, dendritic cells, lymphocytes and hematopoietic stem cells. The following protocol provides a simple and fast method for isolation of bone marrow immune cells (no erythrocytes) from mouse femurs with a yield of approximate 8 x 10⁷ cells in 5 ml culture media (1.6 x 10⁴ cells/µl). Further isolation or flow cytometric analysis might be required for study of specific immune cell types.

Keywords: Blood (血)

Flow cytometry (流cytometry)

Hematopoietic stem cells (造血干细胞)

Materials and Reagents

Sterile paper towel
Sterile surgical pad (Direct Resource, catalog number: 19015742 )
23-gauge (or 25-/26-gauge) needle (BD Biosciences, catalog number: 305145 )
10 ml syringe (BD Biosciences, catalog number: 309604 )
70 µm nylon cell strainer (Falcon, catalog number: 352350 )
Note: Currently, it is “Corning, Falcon^®, catalog number: 11995-065 ”.
50 ml conical tube (Falcon, catalog number: 21008-940 )
Note: Currently, it is “Corning, Falcon^®, catalog number: 21008-940 ”.
5 ml syringe plunger (BD Biosciences, catalog number: 309646 )
Adult mice (> 6 weeks, any strain) (e.g., C57BL/6)
Hank’s balanced salt solution (HBSS), no Calcium, no Magnesium, no phenol red (Life Technologies, catalog number: 14175095 )
Note: Currently, it is “Thermo Fisher Scientific, Gibco^TM, catalog number: 14175095 ”.
DMEM medium, high glucose, pyruvate, L-glutamine (Life Technologies, catalog number: 11995-065 )
Note: Currently, it is “Thermo Fisher Scientific, Gibco^TM, catalog number: 11995-065 ”.
70% ethanol
Fetal bovine serum heat inactivated (FBS) (Sigma-Aldrich, catalog number: F9665 )
Ammonium chloride (NH₄Cl) (Sigma-Aldrich, catalog number: 213330 )
Potassium bicarbonate (KHCO₃) (Sigma-Aldrich, catalog number: 237205 )
Disodium edetate (Sigma-Aldrich, catalog number: D2900000 )
RBC lysis buffer (see Recipes)
DMEM medium (see Recipes)

Equipment

Blunt-end sterile scissors (Thermo Fisher Scientific, Fisher Scientific, catalog number: 08-950 )
Sharp sterile scissors (Thermo Fisher Scientific, Fisher Scientific, catalog number: 08-940 )
Sterile forceps (Thermo Fisher Scientific, Fisher Scientific, catalog number: 08-890 )
Hausser^TM Levy^TM Hemacytometer Chamber Set (Thermo Fisher Scientific, Fisher Scientific, catalog number: 02-671-55A ) or coulter Z2 cell and particle counter (Beckman Coulter, catalog number: 383550 )
Refrigerated centrifuge
Sterile culture hood
CO₂ rodent euthanasia chamber

Procedure

English

中文翻译

文章信息

版权信息

如何引用

Readers should cite both the Bio-protocol article and the original research article where this protocol was used:

Liu, X. and Quan, N. (2015). Immune Cell Isolation from Mouse Femur Bone Marrow. Bio-protocol 5(20): e1631. DOI: 10.21769/BioProtoc.1631.
Liu, X., Yamashita, T., Chen, Q., Belevych, N., McKim, D. B., Tarr, A. J., Coppola, V., Nath, N., Nemeth, D. P., Syed, Z. W., Sheridan, J. F., Godbout, J. P., Zuo, J. and Quan, N. (2015). Interleukin 1 type 1 receptor restore: a genetic mouse model for studying interleukin 1 receptor-mediated effects in specific cell types. J Neurosci 35(7): 2860-2870.