Isolation of Primary Breast Cancer Cells from HER2 Transgenic Mice
从 HER2 转基因小鼠分离原代乳腺癌细胞
发布: 2016年10月05日第6卷第19期 DOI: 10.21769/BioProtoc.1956 浏览次数: 14099
评审: HongLok LungShravani MukherjeeAnonymous reviewer(s)
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参见作者原研究论文
The authors used this protocol in:
Feb 2015
Abstract
HER2 is a tyrosine kinase receptor, which is overexpressed in about 30% of breast cancer patients. Its overexpression leads to mammary tumorigenesis and increased invasion and metastasis (Slamon et al., 1987). HER2 transgenic mouse (FVB/N-MMTVneu mouse) is a well-established model of mammary tumor in human (Fantozzi and Christofori, 2006). Although in vivo models are excellent for assessing the influence of various factors, especially microenvironment, on development of breast cancer, a convenient and less costly way to study the underlying molecular events is utilizing cells derived from the model under evaluation. In order to explore the molecular mechanism by which HOXB7 inhibits initiation, but promotes metastasis of breast tumors, we generated mouse breast cancer cell line from HER2 transgenic mouse (Liu et al., 2015). This protocol may be useful for the generation of breast cancer cell line from mice with other genetic backgrounds.
Keywords: HER2 Transgenic mice (HER2基因的转基因小鼠)
Primary breast cancer cells (原发性乳腺癌细胞)
Mouse breast cancer cell line (小鼠乳腺癌细胞株)
Materials and Reagents
- Cell strainer, sterile (100 μm) (Corning, Falcon®, catalog number: 352360 )
- 10 cm standard tissue culture dish (Corning, catalog number: 430293 )
- 50 ml conical tubes, sterile (Corning, Falcon®, catalog number: 352070 )
- Disposable scalpels, forceps and scissors (VWR International)
- HER2 transgenic mice (THE JACKSON LABORATORY, catalog number: 002376 )
- 0.25% trypsin-EDTA (1x) (Thermo Fisher Scientific, GibcoTM, catalog number: 25200-056 )
- Phosphate-buffered saline (PBS), sterile (self-preparation)
- HEPES (Sigma-Aldrich, catalog number: H3375 )
- DMEM/F12-with L-glutamine (Thermo Fisher Scientific, GibcoTM, catalog number: 11875-093 )
- Bovine serum albumin solution (BSA) (Sigma-Aldrich, catalog number: A9576 )
- Hydrocortisone (Sigma-Aldrich, catalog number: H0888 )
- Type IV collagenase (Sigma-Aldrich, catalog number: C5138 )
- Hyaluronidase (Sigma-Aldrich, catalog number: H3884 )
- Pen-Strep (GE Healthcare, HyCloneTM, catalog number: SV30010 )
- Ammonium chloride (NH4Cl) (Sigma-Aldrich, catalog number: A9434 )
- Tris (Sigma-Aldrich, catalog number: 93362 )
- Fetal bovine serum (FBS), heat inactivated (GE Healthcare, HyCloneTM, catalog number: SH30071.03 )
- Insulin (with transferrin/selenium) (Thermo Fisher Scientific, GibcoTM, catalog number: 51500-056 )
- Digestion buffer (see Recipes)
- Tris-buffered ammonium chloride (TAC buffer) (see Recipes)
- Complete media (see Recipes)
Equipment
- Shaking incubator (Thermo Fisher Scientific, Thermo Scientific, model: SHKA5000 )
- 37 °C, 5% CO2 cell culture incubator (VWR, symphonyTM, model: 5.3A )
- Pipette
- Refrigerated centrifuge (Eppendorf, model: 5418R )
- Water bath
- Hemocytometer
- Inverted microscope (Olympus Corporation, model: CKX41 )
- Tissue culture hood equipped with UV light source (Labconco, model: Purifier Logic+ Class II , Type A2 Biosafety Cabinet)
Procedure
文章信息
版权信息
© 2016 The Authors; exclusive licensee Bio-protocol LLC.
如何引用
Liu, S. and Chen, H. (2016). Isolation of Primary Breast Cancer Cells from HER2 Transgenic Mice. Bio-protocol 6(19): e1956. DOI: 10.21769/BioProtoc.1956.
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分类
癌症生物学 > 癌症干细胞 > 细胞生物学试验
细胞生物学 > 细胞分离和培养 > 细胞分离
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